3.3. Irritability tests
Since the endogenous antioxidant concentration decreases from the outermost to the deeper layers of the skin, natural antioxidants can be applied topically to protect skin from oxidative stress induced by UV radiation, although not all antioxidants have similar affinity for different layers of the skin (Abla and Banga, 2013). In order to confirm the potential of the extracts for topical use, irritability assays with the Episkin test were performed. Data for the cell viability and the cytokine (interleukin-1 , IL-1 ) release from in vitro assays with reconstituted human epidermis are shown in Table 2. This method is based on RhE-models using human keratinocytes, which represent an in vitro model of the target organ for skin irritation and covers the initial step of the inflammatory cascade that occurs during irritation in vivo. Irritant chemicals are identified by their ability to decrease cell viability below defined threshold levels of 50%. The method distinguishes between irritants and
non-irritants. The extracts produced during the autohydrolysis stage and the ethanolic extract from the selected materials are not skin irritants for topical use in cosmetic formulations at 0.1% because the viability in all cases is higher than 50% with values between 79.44 and 92.33%. This potential use was also confirmed by the low release of IL1- as endpoint of the irritation process (in the range of negative control), which is a highly active and pleiotropic pro-inflammatory cytokine playing a key role in inflammation and can be the biological mirror of skin irritation. Moreover