Investigation of methods to screen antioxidant combinations for synergistic protection of skin against UV damage

In recent years, the Australian market has seen an influx in cosmeceutical products that focus on anti-ageing. Antioxidants play a crucial role in combatting the ageing process by protecting against oxidative stress, specifically ultraviolet radiation (UVR) induced. The cosmeceutical industry currently utilises a range of in vitro and in vivo tests to evaluate new compounds and plant extracts for inclusion in anti-ageing products. These tests have assisted formulators in identifying functional bioactive ingredients, and they have provided scientific validation for consumer marketing. However, there has been little exploration of the potential for synergism between multiple antioxidants. The aim of this thesis was to establish an in vitro method for screening combinations of antioxidants for their protective efficacy against UVR induced oxidative stress. An extensive review of relevant scientific literature, patents and industry practice was conducted. This was translated into a structured database of methods divided into four key categories: cellular activity, lipid peroxidation, deoxyribonucleic acid (DNA) alterations, and protein oxidation. These methods range from simple, highly repeatable assays to more complex analyses. Based on this literature review, the MTT assay was selected to test the cellular metabolic activity and assess the antioxidant capacity of various compounds in protecting HaCaT cells from UVR damage. Experimental parameters were investigated to achieve a baseline 50% decrease in viability (LD 50) when cells were exposed to UVR in the absence of antioxidants. When ascorbic acid was included at a concentration of 1 mM or 5 mM, the cell viability was observed to be maintain at to an average of 80% and 90%, respectively, following UVR. Further testing explored potential synergistic effects between ascorbic acid and various other natural and synthetic antioxidants, including tocopherol α, ferulic acid, glutathione (GSH), and γ-glutamylcysteine (GGC). Promising synergism was observed particularly between ascorbic acid and phenolic compounds, where the improvement in cell viability over baseline was greater than the sum of the improvements seen with the individual antioxidants used individually. Antioxidant combinations identified from this screening methodology as having potential synergism should be further explored in more complex testing, including human trials, and may ultimately lead to new formulated anti-ageing cosmeceutical products.