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Microbial-Derived Oils and Value-Added Products: Biosynthesis and Applications for Biofuel Production Utah State University DigitalCommons@USU All Graduate Theses and Dissertations Graduate Studies 5-2015 Microbial-Derived Oils and Value-Added Products: Biosynthesis and Applications for Biofuel Production Alex T. McCurdy Utah State University Follow this and additional works at: https://digitalcommons.usu.edu/etd Part of the Biochemistry Commons Recommended Citation McCurdy, Alex T., "Microbial-Derived Oils and Value-Added Products: Biosynthesis and Applications for Biofuel Production" (2015). All Graduate Theses and Dissertations. 4270. https://digitalcommons.usu.edu/etd/4270 This Dissertation is brought to you for free and open access by the Graduate Studies at DigitalCommons@USU. It has been accepted for inclusion in All Graduate Theses and Dissertations by an authorized administrator of DigitalCommons@USU. For more information, please contact [email protected]. MICROBIAL-DERIVED OILS AND VALUE-ADDED PRODUCTS: BIOSYNTHESIS AND APPLICATIONS FOR BIOFUEL PRODUCTION by Alex T. McCurdy A dissertation submitted in partial fulfillment of the requirements for the degree of DOCTOR OF PHILOSOPHY in Biochemistry Approved: ________________________ _______________________ Lance C. Seefeldt Scott A. Ensign Biochemistry Biochemistry Major Professor Committee Member ________________________ _______________________ Alvan C. Hengge Sean J. Johnson Biochemistry Biochemistry Committee Member Committee Member ________________________ _______________________ Bruce Bugbee Mark R. McLellan Environmental Plant Physiology Vice President for Research and Committee Member Dean of the School of Graduate Studies UTAH STATE UNIVERSITY Logan, Utah 2015 ii Copyright © Alex T. McCurdy 2015 All Rights Reserved iii ABSTRACT Microbial-Derived Oils and Value-Added Products: Biosynthesis and Applications for Biofuel Production by Alex T. McCurdy, Doctor of Philosophy Utah State University, 2015 Major Professor: Dr. Lance C. Seefeldt Department: Chemistry and Biochemistry Efforts are being made to replace petroleum-derived fuels with biofuels in a cost competitive manner. It is apparent that the continued use of petroleum is futile as population and technological growth put increasing pressure on the demand for cheap energy and chemicals. Diminishing resources, civil unrest in the Middle East, and the impact of using petrochemicals on the environment are critical driving forces for research in generating renewable petroleum replacements that can be produced with a limited carbon-footprint. Today, biofuels are derived mostly from land-based plants, but their potential for displacing petroleum is limited due to the competition with available farmland used in food production as well as their relatively slow growth rates. Microorganisms as single-cell factories for the production of biofuels have a promising outlook since they do not compete with the food and feed supply and they lack the necessity of arable land used in cultivation. Viable biofuel production hinges upon iv obtaining sufficient biofuel yields, lowering the costs of processing, production of value- added products, and real-life evaluation of the produced fuels. Herein, the current understanding of microbial biochemistry as well as new findings in the role of ATP citrate lyase in lipid accumulation in oleaginous yeasts will be discussed. Also, description of a novel two-step process that generates biodiesel blends from oleaginous microbes will be given. To facilitate the viability of microbial biofuel production, concomitant production of heterologous lactoferrin using Kluyveromyces lactis will be described. Lastly, the real-life evaluation of these microbial biofuels in a diesel engine is reported. The focus of this dissertation will be directed towards addressing the issues related to microbial biofuel production in order to facilitate the viability of biofuel production such that petroleum use can be displaced by more renewable and clean methodologies. (204 pages) v PUBLIC ABSTRACT Microbial-Derived Oils and Value-Added Products: Biosynthesis and Applications for Biofuel Production by Alex T. McCurdy, Doctor of Philosophy Utah State University, 2015 Major Professor: Dr. Lance C. Seefeldt Department: Chemistry and Biochemistry Efforts are being made to replace petroleum-derived fuels with biofuels in a cost competitive manner. It is apparent that the continued use of petroleum is futile as population and technological growth put increasing pressure on the demand for cheap energy and chemicals. Diminishing resources, civil unrest in the Middle East, and the impact of using petrochemicals on the environment are critical driving forces for research in generating renewable petroleum replacements that can be produced with a limited carbon-footprint. Today, biofuels are derived mostly from land-based plants, but their potential for displacing petroleum is limited due to the competition with available farmland used in food production as well as their relatively slow growth rates. Microorganisms as single-cell factories for the production of biofuels have a promising outlook since they do not compete with the food and feed supply and they lack the necessity of arable land used in cultivation. Viable biofuel production hinges upon vi obtaining sufficient biofuel yields, lowering the costs of processing, production of value- added products, and real-life evaluation of the produced fuels. Herein, the current understanding of microbial biochemistry as well as new findings in the role of ATP citrate lyase in lipid accumulation in oleaginous yeasts will be discussed. Also, description of a novel two-step process that generates biodiesel blends from oleaginous microbes will be given. To facilitate the viability of microbial biofuel production, concomitant production of heterologous lactoferrin using Kluyveromyces lactis will be described. Lastly, the real-life evaluation of these microbial biofuels in a diesel engine is reported. The focus of this dissertation will be directed towards addressing the issues related to microbial biofuel production in order to facilitate the viability of biofuel production such that petroleum use can be displaced by more renewable and clean methodologies. vii ACKNOWLEDGMENTS My time as a graduate student in Biochemistry at Utah State has been excellent. The friendships and professional relationships I have established has made the experience quite pleasant. There is common misconception that graduate school has to be dreadful, but it doesn’t have to be so long as you work on something that you love and you surround yourself with positive and successful people like I have during my time here. I would first like to thank my major professor, Dr. Lance Seefeldt, who has been an exceptional mentor. He has allowed me to work on a project I am passionate about and he has motivated me to be successful. There are a lot of things that makes Dr. Seefeldt great at what he does, but what caught my attention within only a few minutes of meeting him is his ability to draw in an audience and excite people about his research. I hope to take some of the skills I have learned from him and carry them onto the next step in my career. I would like to thank my committee members, Dr. Ensign, Dr. Hengge, Dr. Johnson, and Dr. Bugbee for taking the time to provide guidance and helping me develop as a scientist. I thank my lab associates Brad, Danyal, Zhiyong, Robert, Valerie, Rhesa, Sudipta, Nimesh, AJ, Andy, Mike, Tawnie, and Derek for providing an inquisitive and fun atmosphere to work in. It has also been great to work with all the other graduate students in the Chemistry and Biochemistry department while I have been here as well. I would also like to thank our various collaborators, specifically Dr. Jason Quinn and Hailey Summers. viii Most importantly, I have to thank my family. I cannot thank my father Alan and my mother Kim enough for believing in me and supporting me. My wife, Ashley, has made all this possible as she cared for and nurtured our son Jaxton and me. I am forever grateful for her love and support, and I am excited to see what the future holds for us. Alexander Trent McCurdy ix I dedicate this dissertation to my son, Jaxton. x CONTENTS Page ABSTRACT ....................................................................................................................... iii PUBLIC ABSTRACT .........................................................................................................v ACKNOWLDEGMENTS ................................................................................................ vii LIST OF TABLES ............................................................................................................ xii LIST OF FIGURES ......................................................................................................... xiii ABBREVIATIONS ...........................................................................................................xv CHAPTER 1. INTRODUCTION .............................................................................................1 Extraction, Processing, and Purification of Triglyceride Oils .....................6 Microbial Biofuel .........................................................................................8 Lipid Biochemistry ....................................................................................18 Value-Added Products ...............................................................................26 References ..................................................................................................32
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