Optimization of rutin isolation from Amaranthus paniculatus leaves by high pressure extraction and fractionation techniques

A procedure based on the application of pressurized liquid extraction (PLE) was employed to recover the major Amaranthus spp. flavonoid glycoside rutin from dried ground plant leaves. A central composite design (CCD) was used to optimize rutin extraction; three factors were considered: extraction solvent (water/ethanol ratio, from 0/100 to 100/0), extraction time (from 5 to 20 min) and extraction temperature (from 50 to 200 °C). Optimization of the three extraction parameters revealed that water/ethanol ratio in the solvent mixture was the most-significant factor influencing rutin yield, followed by extraction temperature. The total rutin recovery varied from 6.95 to 14.03 g/kg dry matter (DM) depending on the experimental conditions tested. The optimum extraction conditions for the highest rutin content were 70/30 (v/v) water/ethanol ratio, at 188 °C and 20 min as static extraction time. In this case rutin recovery from the leaves (14.30 g/kg DM) was 4-times higher compared to that attainable using conventional ultrasound assisted extraction (3.62 g/kg DM) with methanol:ethanol mixture (90:10) as solvent. Rutin recovery from defatted and non-defatted seeds of amaranth under optimum PLE conditions was 35.3 and 41.1 mg/kg DM, respectively, whereas using ultrasound assisted extraction 19.7 and 34.6 mg/kg DM were obtained, respectively. Extracts obtained under optimum conditions were further fractionated by using supercritical antisolvent fractionation (SAF). Extracts of amaranth leaves containing 70% water were mixed with supercritical CO2 and separated into two fractions according to the molecular mass and solubility of the contained compounds. A central composite experimental design was applied for the determination of the best fractionation conditions to maximize rutin enrichment. Maximum rutin concentration in raffinate (22.57 g/kg) was recovered at 15 MPa using a feed mixture flow rate of 0.3 mL/min.The financial support for the scientific mission at the Institute of Food Science Research (CIAL-CSIC) was funded by the EU Structural Funds project “PhD Students Internship to the Overseas Research Centres” within the framework of the Measure for Enhancing Mobility of Scholars and Other Researchers and the Promotion of Student Research (VP1-3.1-ŠMM-01-V) of the Programme of Human Resources Development Action Plan. M.H. thanks MICINN for his “Ramón y Cajal” research contract.Peer Reviewe