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University of Florida Thesis Or Dissertation Formatting THE EFFECTS OF SOIL TYPE AND NITROGEN RATES ON GLUCOSINOLATE AND OIL PRODUCTION IN BRASSICA CARNATA By THEODOR LINARES STANSLY A THESIS PRESENTED TO THE GRADUATE SCHOOL OF THE UNIVERSITY OF FLORIDA IN PARTIAL FULFILLMENT OF THE REQUIREMENTS FOR THE DEGREE OF MASTER OF SCIENCE UNIVERSITY OF FLORIDA 2016 © 2016 Theodor Linares Stansly To my parents, Philip and Silvia Stansly, I thank you from the bottom of my heart. Not all of the important lessons needed for a successful career can be taught in schools or universities. They must also come from those who have persevered through life’s obstacles without losing hope, inspiration, and spirit. I am grateful for everything you have done and continue to do throughout these years. ACKNOWLEDGMENTS Many thanks to Pete C. Andersen for allowing me the opportunity to pursue my passion for science and providing the steppingstones to a career as a professional researcher. I also want to thank David Wright, Jim Marois, Sheeja George, and Ramdeo (Andy) Seapaul and the rest of the Brassica carinata research team at the UF North Florida Research and Education Center for all your support and assistance in the development of my project. Mosbah M. Kushad from the University of Illinois and Steven H. Miller at the University of South Florida for assisting in various aspects of data acquisition of the samples. 4 TABLE OF CONTENTS page ACKNOWLEDGMENTS .................................................................................................. 4 LIST OF TABLES ............................................................................................................ 7 LIST OF FIGURES .......................................................................................................... 8 LIST OF ABBREVIATIONS ............................................................................................. 9 ABSTRACT ................................................................................................................... 10 CHAPTER 1 LITERATURE REVIEW .......................................................................................... 12 Climate Change and its Origins .............................................................................. 12 Renewable Fuel Standard ...................................................................................... 13 Long Chain Fatty Acids ........................................................................................... 14 Brassica carinata: The “New” Competitor ............................................................... 16 The Glucosinolate-Myrosinase Defense System .................................................... 19 2 THE EFFECT OF SOIL TYPE ON GLUCOSINOLATE PRODUCTION IN BRASSICA CARINATA DURING VARIOUS STAGES OF ITS GROWTH CYCLE .................................................................................................................... 25 Introduction ............................................................................................................. 25 Materials and Methods ............................................................................................ 27 Greenhouse and Irrigation ................................................................................ 27 Soil ................................................................................................................... 27 Tissue Sampling ............................................................................................... 28 Glucosinolate Extraction and Purification ......................................................... 28 Glucosinolate Quantification ............................................................................. 29 Glucosinolate Profiles ....................................................................................... 30 Experimental Design and Statistics .................................................................. 31 Results .................................................................................................................... 31 Roots ................................................................................................................ 31 Stem ................................................................................................................. 32 Leaves .............................................................................................................. 32 Seed ................................................................................................................. 33 Discussion .............................................................................................................. 33 3 THE INFLUENCE OF NITROGEN RATES ON OIL AND GLUCOSINOLATES IN SEEDS OF BRASSICA CARINATA ................................................................... 39 Introduction ............................................................................................................. 39 5 Materials and Methods ............................................................................................ 40 Soil ................................................................................................................... 40 Irrigation and Fertilization ................................................................................. 41 Glucosinolate and Fatty Acid Profiles ............................................................... 42 Experimental Design and Statistics .................................................................. 42 Results .................................................................................................................... 43 Discussion .............................................................................................................. 44 4 COMPARING TWO ETHIOPIAN MUSTARD GENOTYPES GROWN IN THREE TYPES OF AGRICULTURALLY MANAGED SOILS .............................................. 47 Introduction ............................................................................................................. 47 Materials and Methods ............................................................................................ 48 Seed and Soil ................................................................................................... 48 Irrigation and Fertilization ................................................................................. 49 Glucosinolate and Fatty Acid Profiles ............................................................... 50 Experimental Design and Statistics .................................................................. 50 Results .................................................................................................................... 51 Discussion .............................................................................................................. 51 5 CONCLUSION ........................................................................................................ 55 Volatile Identification and Quantification ................................................................. 55 Myrosinase Assays ................................................................................................. 55 Prospecting for Carinata’s Future ........................................................................... 57 LIST OF REFERENCES ............................................................................................... 59 BIOGRAPHICAL SKETCH ............................................................................................ 65 6 LIST OF TABLES Table page 2-1 The pH, nutrients (ppm), organic matter (%) and electrical conductivity (EC) of the three soil types for potting B. carinata in a greenhouse. ........................... 36 2-2 The Hoagland’s nutrient solution at varying nitrogen rates. ................................ 36 2-3 Dry biomass of root, leaf, stem, seed of Brassica carinata grown in three soil types. .................................................................................................................. 36 2-4 Total Glucosinolate and oil content in the seeds of conventionally grown carinata using near infrared spectroscopy. ......................................................... 37 7 LIST OF FIGURES Figure page 1-1 Concept map for producing biodiesel via transesterification. .............................. 22 1-2 Flow chart for the conversion of triglycerides into biojet or biodiesel via catalytic hydrothermolysis, hydroprocessing, and fractionation. ......................... 22 1-3 Comparison of long chain fatty acid profiles from various oilseed crops as percent (%) of total oil. Source: .......................................................................... 23 1-4 Diagram of possible outcomes of glucosinolate and its derivative (isothiocyanate) after soil incorporation. ............................................................. 24 2-1 Total glucosinolate concentration of the roots, stem, and leaves in Brassica carinata at different stages in its life cycle. ......................................................... 38 3-1 Total biomass accumulation at various rates of nitrogen using modified versions of Hoagland solutions with standard error ............................................ 46 3-2 Distribution of oil, protein, total glucosinolate concentration, erucic acid, long and very long chain fatty acid in the seed, .......................................................... 46 4-1 Percentage of seeds germination and establishment. ........................................ 53 4-2 Correlation
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