The present study concerns the “in vitro” safety evaluation, using human cells. Cytotoxicity through the NRU assay has been determined in order to assess product safety. NRU assay is “in vitro” evaluation of the cytotoxicity potential of cosmetic products. Neutral red uptake assay for the estimation of cell viability/cytotoxicity

The neutral red uptake assay provides a quantitative estimation of the number of viable cells in a culture. It is one of the most used cytotoxicity tests and more sensitive than other cytotoxicity tests with many biomedical and environmental applications. It is based on the ability of viable cells to incorporate and bind the supravital dye neutral red in the lysosomes.

In vitro methods are an interesting alternative system to traditional in vivo tests to evaluate biological properties of cosmetic ingredients and products, according to the current European cosmetic rules that ask manufacturers to assess the product safety, without employing animals (Basic Council Directive N° 76/768/ EEC of the 27/07/76, EC L. 262 of the 27/09/1976; VI Amendment Council Directive 93/35 EEC of the 14/06/1993 ECL.151 of the 23/06/1993).

The “in vitro” test on derma-derived cells is a simplified but yet very informative model of the reactions that may occur “in vivo”. Cosmetic products are directly applied locally on the human skin, often in contact with mucous membranes. Consequently they should exhibit no or very low toxicity to the epithelial cells. Cytotoxicity test performed on human keratinocytes or fibroblasts in monolayer have been extensively used as indicator of the eye and skin irritation potential of cosmetics and ingredients.