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Center for Food Science & Nutrition ADDIS ABABA UNIVERSITY COLLEGE OF NATURAL SCIENCES CENTER FOR FOOD SCIENCE AND NUTRITION STUDIES ON PHENOLIC CONTENTS AND BIOFUNCTIONAL ACTIVITIES OF SELECTED ETHIOPIAN SPICES AND HERBS By Engeda Dessalegn Dissertation Submitted to the Center for Food Science and Nutrition in Partial Fulfillment of the Requirements for the Degree of Doctor of Philosophy Supervisors: Dr. Geremew Bultosa (Associate Professer. Haramaya University, Ethiopia) Prof. Gulelat Dessie Haki (Addis Ababa University, Ethiopia) Dr. H. P. Vassantha Rupasinghe (Associate Professer, Dalhousie University, Agricultural College, Canada) June, 2015 Addis Ababa Center for Food Science & Nutrition The undersigned examining committee approved the thesis entitled: Studies on Phenolic contents and Biofunctional Activities of Selected Ethiopian Spices and Herbs A. Examiners 1. Professor Gun-Hee Kim __________________________________________________ 2. Dr Daniel Bisrsat (Associate Professer) ____________________________________________________ B. Chairperson Ato Tilahun Bekele (Assistant Professor) _________________________________________________________ C. Advisers 1. Dr. Geremew Bultosa (Associate Professor) __________________________________________________________ 2. Professer Gulelat Desse Haki ___________________________________________________________ 3. Dr. Vassantha Rupasinghe (Associate Professor) _____________________________________________________________ June 29, 2015 Addis Ababa, Ethiopia PhD dissertation Page ii Center for Food Science & Nutrition Dedication This dissertation is dedicated for those eighty four civilians, including my Dad, who were beheaded and shot down by Somali invaders in a place called Biyo Keraba, Doba Woreda, West Harrarghe (Eastern Ethiopia), on June 30, 1977 G.C. May your souls continue to rest in peace? PhD dissertation Page iii Center for Food Science & Nutrition TABLE OF CONTENTS LIST OF FIGURES………………………………………………………………………………ix LIST OF TABLES……………………………………………………………………………...xi ii ACKNOWLEDGEMENT…………………………………………………………………….....xv ACRONYMS…..……………………………………………………………………………………………………………………………….xvii ABSTRACT…………………………………………………………………………………………………………………………………….,.xx CHAPTER 1: GENERAL BACKGROUND ............................................................................. 1 1.1 General Introduction ................................................................................................................. 1 1.2 Literature Review...................................................................................................................... 4 1.2.1 Dietary spices and herbs ..................................................................................................... 4 1.2.2 Dietary spices and herbs in Ethiopia .................................................................................. 5 1.2.3 General description of selected dietary spice and herb plants used for the study .............. 6 1.2.3.1 Aframomum corrorima ................................................................................................ 6 1.2.3.2 Coriandrum sativum .................................................................................................... 7 1.2.3.3 Lippia adoensis ............................................................................................................ 9 1.2.3.4 Thymus schimperi Ronniger ...................................................................................... 11 1.2.4 Introduction to phenolic phytochemicals ......................................................................... 12 1.2.4.1 Phenolic compounds .................................................................................................. 13 1.3 Statement of the Problem…………………………………………………………………….26 1.4. Objectives .............................................................................................................................. 28 1.4.1. General objective............................................................................................................. 28 1.4.2 Specific objectives............................................................................................................ 29 CHAPTER 2: EVALUATION OF PHENOLIC COMPOSITION OF Aframomum corrorima, Thymus schimperi, Lippia adoensis, AND Coriandrum sativum ............................. 30 PhD dissertation Page iv Center for Food Science & Nutrition Abstract ......................................................................................................................................... 30 2.1. Introduction ............................................................................................................................ 31 2.2. Materials and Methods ........................................................................................................... 34 2.2.1. Standards and chemicals ................................................................................................. 34 2.2.2. Plant materials ................................................................................................................. 34 2.2.3 Preparation of plant extracts for the determination of total phenolic and flavonoid contents...................................................................................................................................... 35 2.2.4. Sample extraction for LC-MS analysis ........................................................................... 35 2.2.5. Determination of total phenolic content (TPC) ............................................................... 36 2.2.6. Determination of total flavonoid content (TFC) ............................................................. 36 2.2.7 Identification of phenolic compounds .............................................................................. 38 2.2.8. Quantification of phenolic compounds ........................................................................... 38 2.3 Result ...................................................................................................................................... 40 2.3.1Total phenolic content ....................................................................................................... 40 2.3.2 Total flavonoid content .................................................................................................... 43 2.3.3 Characterization of phenolic compounds ......................................................................... 46 2.3.4 Quantification of the phenolic compounds ...................................................................... 46 2.3.4.1 Phenolic acids ............................................................................................................ 52 2.3.4.2 Flavonols ................................................................................................................... 53 2.3.4.3 Flavan-3-ols ............................................................................................................... 55 2.3.4.4 Aliphatic organic acids and diydrochalcones ............................................................ 55 Conclusions ................................................................................................................................... 56 CHAPTER 3: IN VITRO ANTIOXIDANT ACTIVITIES OF Aframomum corrorima, Thymus schimperi, Lippia adoensis, AND Coriandrum sativum .............................................. 57 Abstract ......................................................................................................................................... 57 3.1. Introduction ............................................................................................................................ 58 3.2. Materials and Methods ........................................................................................................... 62 3.2.1. Chemicals ........................................................................................................................ 62 PhD dissertation Page v Center for Food Science & Nutrition 3.2.2. Preparation and extraction of plant materials .................................................................. 62 3.2.3. Determination of antioxidant activity ............................................................................. 63 3.2.3.1 DPPH method ............................................................................................................ 63 3.2.3.2 Ferric ion reducing power .......................................................................................... 64 3.2.3.3 Total antioxidant capacity using phosphomolybdenum assay .................................. 66 3.2.3.4 Chelating effects on ferrous ions ............................................................................... 67 3.2.3.5 Extraction of flaxseed oil ........................................................................................... 68 3.2.3.6 Inhibitory activity toward lipid peroxidation (FTC assay) ........................................ 68 3.2.3.7 Thiobarbituric acid reactive substances (TBARS) assay .......................................... 69 3.2.4 Statistical analysis ............................................................................................................ 70 3.3 Results ..................................................................................................................................... 71 3.3.1 In vitro antioxidant activity .............................................................................................
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