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Optimisation of Fractionation Process of Amaranth (Amaranthus Spp.) by Applying High Pressure Extraction Methods

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Optimisation of Fractionation Process of Amaranth (Amaranthus Spp.) by Applying High Pressure Extraction Methods KAUNAS UNVERSITY OF TECHNOLOGY PAULIUS KRAUJALIS OPTIMISATION OF FRACTIONATION PROCESS OF AMARANTH (AMARANTHUS SPP.) BY APPLYING HIGH PRESSURE EXTRACTION METHODS Doctoral Dissertation Technological Sciences, Chemical Engineering (05T) 2014, Kaunas The research was carried out at Kaunas University of Technology (Lithuania), Faculty of Chemical Technology, Department of Food Science and Technology in the period of 2010-2014. The research was in part supported by European funds. Scientific Supervisor: Prof. dr. Petras Rimantas Venskutonis (Kaunas University of Technology, Technological Sciences, Chemical Engineering, 05T). © P. Kraujalis, 2014 KAUNO TECHNOLOGIJOS UNIVERSITETAS PAULIUS KRAUJALIS BURNOČIO (AMARANTHUS SPP.) FRAKCIONAVIMO PROCESŲ OPTIMIZAVIMAS TAIKANT DIDELIO SLĖGIO EKSTRAKCIJOS BŪDUS Daktaro disertacija Technologijos mokslai, chemijos inžinerija (05T) 2014, Kaunas Disertacija parengta 2010-2014 metais Kauno technologijos universitete, Cheminės technologijos fakultete, Maisto mokslo ir technologijos katedroje. Tyrimai buvo iš dalies finansuoti Europos sąjungos struktūrinių fondų lėšomis. Mokslinis vadovas: Prof. dr. Petras Rimantas Venskutonis (Kauno technologijos universitetas, technologijos mokslai, chemijos inžinerija, 05T). © P. Kraujalis, 2014 ACKNOWLEDGEMENTS I am using this opportunity to express my gratitude to everyone who supported me throughout the path of this thesis. I would like to express my gratitude for my scientific promoter Prof. dr. Petras Rimantas Venskutonis for academic guidance during preparation of this dissertation and publications, for the subject of this doctoral dissertation, for continuous encouragement. I am greatly thankful to him as he was available for my support and advice in the time of need and on a number of issues related to this thesis. Prof. dr Elena Ibáñez (Institute of Food Science Research) for the internship in her scientific group as well as friendly atmosphere and valuable scientific advices. I am grateful to all colleagues from the Department of Food Science and Technology – who provide me with the facilities being required, for their friendly advice and help during the experiments as well as friendly environment. I express my deepest affection, appreciation and thanks to my colleague, best friend and beloved wife Vaida and my daughters Elzė and Morta for their love, patience, keeping me on the right track and understanding during difficult moments and also shared my success. Very important thanks to my mother, sister and brother, for their moral support, invaluable guidance, illuminating view and constructive criticism. Also big thanks to all my relatives and friends. I would like to thanks to everyone who contributed to this dissertation. Paulius Kraujalis TABLE OF CONTENT 1. LIST OF ABBREVIATIONS .......................................................................... 7 I. INTRODUCTION ............................................................................................ 9 II. LITERATURE REVIEW ............................................................................... 12 2.1. Supercritical fluid extraction ................................................................... 12 2.2. Others pressurised fluid techniques ......................................................... 30 2.3 Process optimisation methodology ........................................................... 34 2.4. Amaranth ................................................................................................. 35 2.5. Deduction ................................................................................................ 43 III. MATERIALS AND METHODS ................................................................... 44 3.1. Accelerated solvent extraction of lipids from Amaranthus spp. seeds and characterization of their composition ................................................................... 44 3.2. Supercritical carbon dioxide extraction of amaranth seeds by response surface methodology and characterization of extracts ......................................... 46 3.3. Supercritical carbon dioxide extraction of squalene and tocopherols from amaranth and assessment of extracts antioxidant activity .................................... 48 3.4. Antioxidant properties and preliminary evaluation of phytochemical composition of different anatomical parts of amaranth ........................................ 53 3.5. Optimisation of rutin isolation from different anatomical parts of Amaranthus paniculatus using accelerated extraction with green solvents ......... 57 IV. RESULTS AND DISCUSION ....................................................................... 61 4.1. Accelerated solvent extraction of lipids from Amaranthus spp. seeds and characterization of their composition ................................................................... 61 4.2. Supercritical carbon dioxide extraction of amaranth seeds by response surface methodology and characterization of extracts ......................................... 69 4.3. Supercritical carbon dioxide extraction of squalene and tocopherols from amaranth and assessment of extracts antioxidant activity .................................... 76 4.4. Antioxidant properties and preliminary evaluation of phytochemical composition of different anatomical parts of amaranth ........................................ 85 4.5. Rutin isolation from different anatomical parts of Amaranthus paniculatus using accelerated extraction with green solvents .............................. 91 4.6 Bio-refinery of amaranth anatomical parts ............................................... 98 V. CONCLUSIONS .......................................................................................... 101 VI. LITERATURE REFERENCES ................................................................... 103 VII. LIST OF PUBLICATIONS ON THE THEME OF THIS DISERTATION 117 1. LIST OF ABBREVIATIONS ABTS●+ 2,2’-azinobis-3-ethylbenzothiazoline-6-sulfonic acid radical cation ANOVA Analysis of variance AS Analytical scale AscA Ascorbic acid ASE Accelerated solvent extraction CCD Central composite design CERP Constant extraction rate period DCRP Diffusion controlled rate period DPPH● 2,2-diphenyl-1-picrylhydrazyl radical FA Fatty acid FAME Fatty acid methyl ester FERP Falling extraction rate period FID Flame ionisation detector FRAP Ferric reducing antioxidant power GAE Gallic acid equivalents GC Gas chromatography GRAS Generally recognized as safe HPLC High performance liquid chromatography IDF Insoluble dietary fibre LC Liquid chromatography MCERP Mass-transfer rate during CERP MS Mass spectrometry MSD Mass selective detector OEC Overall extraction curve OEC Overall extraction curve PAHs Polycyclic aromatic hydrocarbons PHWE Pressurised hot water extraction PLE Pressurised liquid extraction PS Pilot scale PUFA Poly unsaturated fatty acids QTOF Quadrupole time-of-flight RCERP Yield of extract during CERP RSC Radical scavenging capacity RSD Relative standard deviation RSM Response surface methodology SAF Supercritical antisolvent fractionation SC-CO2 Supercritical carbon dioxide SC-CO2 Supercritical carbon dioxide SCE-CO2 Supercritical extraction carbon dioxide SDF Soluble dietary fibre SFE Supercritical fluid extraction SL Standard litre at standard state (PCO2=100 kPa, TCO2=20 °C, ρCO2=0.0018 g/ml) 7 SWE Subcritical water extraction tCERP Time of CERP TDF Total dietary fibre TEAC Trolox equivalent antioxidant capacity UPLC Ultra high performance liquid chromatography 8 I. INTRODUCTION Relevance of the research. The search for bioactive compounds of natural origin has arisen in the past two decades. Consideration of raw material as well as environmentally-friendly process application is of importance. In last decades, supercritical fluid technology has gained much interest in pharmaceutical and food sectors because of an increased preference for natural products and regulations related to toxicity levels of the natural bioactive substances. Natural new generation products without residues of the chemicals have made supercritical fluid technology as an alternative to traditional solvent extraction for the extraction and fractionation of active ingredients. Different sources of bioactive compounds have been studied; plants, agricultural by-products and marine products being among the most promising, some have already been associated with lower risks of coronary heart diseases and cancer (Hooper, et al., 2008). The supercritical carbon dioxide (SC-CO2) as a solvent has been of choice for food applications. Advantage of processing with SC-CO2 include: low processing temperatures, minimal thermal degradation of the bioactive components, ease of separation of extraction solvent, no solvent residue left in the product, processing in the CO2 environment minimizes undesirable oxidation reactions, which is especially beneficial for the sensitive bioactive components of specialty oils such as sterols, tocols, carotenoids, and polyunsaturated fatty acids (PUFAs) (Temelli, et al., 2007). SCE-CO2 was successfully applied as a separation and fractionation technique for reducing free fatty acid content, minimizing phytosterols loss (Dunford & King, 2000) and for obtaining the fractions enriched with tocopherols and tocotrienols (Sarmento, et al., 2006). The genus Amaranthus includes approximately 60 species growing in many areas of the world. Amaranth has been consumed for centuries as a green leafy vegetable, grain or used for ornamental purposes. Amaranthus is a gluten-free pseudocereal,
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