Biological activity of ethanol extract from leaves of Rosmarinus eriocalyx

Aging or senescence is a complex and inevitable process, which is not only attributed to individual genetic variation but also to external factors such as environmental conditions, nutrition, alcohol, and diseases [1]. The most widely accepted theory, that have been proposed to explain aging, is the free radical theory [2]. Aging and related diseases result from accumulated oxidative damage to cell constituents and tissues caused by excessive exposure to free radicals. Antioxidants, which mediate the imbalance between intracellular antioxidant defenses and oxidative damage by reducing the reactive oxygen species (ROS) levels, are believe to be able to reduce stress-induced premature senescence or slow down replicative senescence [3]. Rosmarinus eriocalyx (Jord. & Fourr.) is an aromatic evergreen bush belonging to Lamiaceae family and endemic to Algeria, Morocco and Spain, where it is used as a condiment to flavor soup and meat and as a traditional remedy [4]. The plant volatile fraction is characterized by the monoterpene ketone camphor, whereas its ethanolic extracts are rich sources of phenolic acids and diterpenes such as rosmarinic acid, carnosic acid and carnosol that are the main responsible for the noteworthy antioxidant activity [5]. In this setting, we aimed to evaluate R. eriocalyx biological activity in order to propose the plant as an anti-aging agent. For this purpose, we determined the cytotoxic activity of polar extracts obtained from leaves, flowers, and stems of R. eriocalyx on human fibroblast and human tumor cell lines (A375, MDA-MB 231, and T98G) by MTT assay [6]. Results showed that the ethanolic extract of leaves resulted the most active against A375 human melanoma cell line (IC50 value of 17.8 µg/ml). The total phenolic content values reported for R. eriocalyx ethanolic and aqueous extracts showed slight differences and free radical scavenging activity was stronger for ethanolic extracts than aqueous ones. On this basis, we selected the R. eriocalyx ethanolic extract to determine the antioxidant activity on human fibroblast by measuring its ability to prevent oxidation in cells using a ROS fluorescent probe (DCFH-DA) [7]. Results showed a remarkable activity in preventing oxidation of cells induced by 2.2’-azobis -2-amidinopropane (ABAP). Afterwards, we tested the same extract on the H2O2-induced premature senescence in young fibroblast cells where -galactosidase (SA--gal) activity was used to measure cell senescence [8]. Preliminary data showed a reduction of H2O2 stress-induced premature senescence indicating the potential of R. eriocalyx leaf extract to be formulated as an anti-aging agent. References [1] N. Getoff. Anti-aging and aging factors in life. The role of free radicals. Radiat. Phys. Chem. 2007, 76,1577-1586. [2] D. Harman. Aging: a theory based on free radical and radiation chemistry. J. Gerontol. 1956, 11, 298-300. [3] D. Fusco, G. Colloca, M.R. Lo Monaco, M. Cesari. Effects of antioxidant supplementation on the aging process. Clin. Interv. Aging. 2007, 2, 377-387. [4] M.S. Bendeddouche, H. Benhassaini, Z. Hazem, A. Romane. Essential oil analysis and antibacterial activity of Rosmarinus tournefortii from Algeria. Nat. Prod. Commun. 2011, 6, 1511-1514. [5] H. Bendif, M. Boudjeniba, M. Djamel Miara, L. Biqiku, M. Bramucci, G. Caprioli, G. Lupidi, L. Quassinti, G. Sagratini, L.A. Vitali, S. Vittori, F. Maggi. Rosmarinus eriocalyx: An alternative to Rosmarinus officinalis as a source of antioxidant compounds. Food Chem. 2017, 218, 78-88. [6] L. Quassinti, G. Lupidi, F. Maggi, F. Papa, S. Vittori, A. Bianco, M. Bramucci . Antioxidant and antiproliferative activity of Hypericum hircinum L. subsp. majus (Aiton) N. Robson essential oil. Nat. Prod. Res. 2013, 27, 862-868. [7] K.L. Wolfe, R.H. Liu. Cellular antioxidant activity (CAA) assay for assessing antioxidants, foods, and dietary supplements. J. Agric. Food Chem. 2007, 55, 8896-8907. [8] D.J. Kurz, S. Decary, Y. Hong, J.D. Erusalimsky. Senescence-associated -galactosidase reflects an increase in lysosomal mass during replicative ageing of human endothelial cells. J. Cell Sci. 2000, 113, 3613–3622.