Supercritical antisolvent fractionation of rosemary extracts obtained by pressurized liquid extraction to enhance their antiproliferative activity

Rosemary bioactivity is correlated with the presence of phenolic diterpenes (carnosic acid, carnosol) and phenolic acids (rosmarinic acid). In the present work, two processes, pressurized liquid extraction (PLE) and supercritical antisolvent fractionation (SAF) were integrated to separate and enrich these families of compounds. The optimization of the SAF process was carried out using a response surface methodology (RSM) with three factors: (i) CO pressure, (ii) percentage of water in the PLE extract and (iii) PLE extract/SC-CO flow ratio. The selected responses were the relative amount of each compound (rosmarinic acid (RA) and carnosic acid + carnosol (CA + CS)), total phenolic content, antioxidant activity and antiproliferative activity against HT-29 colon cancer cells. The higher CA + CS content was obtained at 100 bar, 50% (v/v) of water in the feed solution and 0.025 feed/SC-CO mass flow ratio, achieving values of 478.1 mg/g extract and 17% of cell survival after 24 h of treatment.A.P.S.C. thanks to the Administrative Department of Science, Technology and Innovation COLCIENCIAS (Colombia) for her Ph.D. Scholarship. A.V. thanks the Ministerio de Economía y Competitividad for his FPI pre-doctoral fellowship. M.H. would like to thank MICINN for his “Ramón y Cajal” research contract. The authors would also like to thank Projects AGL2011-29857-C03-01, AGL2014-53609-P (MINECO, Spain) and ALIBIRD-CM, S2013/ABI-2728 (Comunidad de Madrid) for their financial support.Peer Reviewe