Evaluation of chemopreventive activity of spinach extracts of plant grown in normal and hypoxic condition

Consumption of fresh fruits and vegetables in the human diet is important to maintain a good health status and to prevent chronic diseases. It is well known that plant derived food is a powerful source of chemopreventive molecules such as antioxidants (polyphenols, catechins, etc..). Stress response represents a powerful stimulus for plants to produce metabolites with high value for human health. However, to date, this approach has not been extensively used yet, since too much mechanistic information is still lacking. To find responses to this, we have investigated Spinach (Spinacia oleracea L., Chenopodiaceae) grown in normal or in hypoxic condition. Spinach leaves, commonly present in the human diet both raw and cooked, are a concentrate of bio-active compounds, molecules with a great medical value but rarely diffused in the Plant Kingdom such as phytoecdysteroids. The aqueous extracts of lyophilized Spinach leaves have been administered to tumoral colon cell line HT-29 to evaluate their chemoprotective effects. Cell viability assay (MTS) have been used to assess the antiproliferative activity while Comet Assay have been used to consider the cito-genotoxity of the extracts. The extracts of plants grown in hypoxic condition exert an antiproliferative activity greater than those grown in normal conditions. None of the extracts exerted any genotoxic activity when tested alone. To better understand the way of action of the antiproliferative activity we found, we tested the antioxidant activity of the extracts by the comet assay, after a co-treatment with a known oxidizing agent as H2O2, and the induction of apoptosis in HT-29 cell line by TUNEL assay. Unexpectedly, the spinach extracts did not shown any antioxidants activity in vitro, meanwhile they seemed to induce apoptosis. The analyses of the chemical composition of the extracts are ongoing, they could permit us to identify the chemical mixture present in the different extracts and which compound is responsible for the increased antiproliferative activity shown by the plants grown in hypoxia. What we found does not confirm what reported by Moser that showed an antioxidant activity of spinach in in vivo studies. Taking into account these observations, we plan to simulate the digestion of the extracts to understand if the antioxidant activity could be due to a chemical modification of the original mixture