Mississippi State University Scholars Junction Theses and Dissertations Theses and Dissertations 1-1-2010 Enhanced Lipid Production And Biodiesel Yields From Activated Sludge Via Fermentation Of Lignocellulose Hydrolyzate Andro Hernandez Mondala Follow this and additional works at: https://scholarsjunction.msstate.edu/td Recommended Citation Mondala, Andro Hernandez, "Enhanced Lipid Production And Biodiesel Yields From Activated Sludge Via Fermentation Of Lignocellulose Hydrolyzate" (2010). Theses and Dissertations. 1887. https://scholarsjunction.msstate.edu/td/1887 This Dissertation - Open Access is brought to you for free and open access by the Theses and Dissertations at Scholars Junction. It has been accepted for inclusion in Theses and Dissertations by an authorized administrator of Scholars Junction. For more information, please contact [email protected]. Template Created By: Damen Peterson 2009 ENHANCED LIPID PRODUCTION AND BIODIESEL YIELDS FROM ACTIVATED SLUDGE VIA FERMENTATION OF LIGNOCELLULOSE HYDROLYZATE By Andro Hernandez Mondala A Dissertation Submitted to the Faculty of Mississippi State University in Partial Fulfillment of the Requirements for the Degree of Doctor of Philosophy in Engineering in the Dave C. Swalm School of Chemical Engineering Mississippi State, Mississippi December 2010 Template Created By: Damen Peterson 2009 Copyright by Andro Hernandez Mondala 2010 Template Created By: Damen Peterson 2009 ENHANCED LIPID PRODUCTION AND BIODIESEL YIELDS FROM ACTIVATED SLUDGE VIA FERMENTATION OF LIGNOCELLULOSE HYDROLYZATE By Andro Hernandez Mondala Approved: __________________________________ __________________________________ Rafael Hernandez W. Todd French Associate Professor Assistant Professor Dave C. Swalm School of Dave C. Swalm School of Chemical Engineering Chemical Engineering (Director of Dissertation and Advisor) (Committee Member) __________________________________ __________________________________ R. Mark Bricka Bill Elmore Associate Professor Interim Director and Henry Hunter Chair Dave C. Swalm School of Dave C. Swalm School of Chemical Engineering Chemical Engineering (Committee Member) (Committee Member) __________________________________ __________________________________ Benjamin S. Magbanua, Jr. Keisha B. Walters Associate Professor Assistant Professor Department of Civil and Dave C. Swalm School of Environmental Engineering Chemical Engineering (Committee Member) (Committee Member) __________________________________ __________________________________ Rafael Hernandez Sarah A. Rajala Associate Professor Dean of the Bagley College of Engineering Dave C. Swalm School of Chemical Engineering (Graduate Coordinator) Template Created By: Damen Peterson 2009 Name: Andro Hernandez Mondala Date of Degree: December 10, 2010 Institution: Mississippi State University Major Field: Engineering Major Professor: Rafael Hernandez, Ph.D. Title of Study: ENHANCED LIPID PRODUCTION AND BIODIESEL YIELDS FROM ACTIVATED SLUDGE VIA FERMENTATION OF LIGNOCELLULOSE HYDROLYZATE Pages in Study: 316 Candidate for Degree of Doctor of Philosophy The potential of enhancing lipid accumulation in municipal sewage activated sludge via fermentation of lignocellulose biomass hydrolyzate was investigated. The overall objective was to increase the levels of feedstock lipids in the activated sludge biomass and increase its biodiesel yield via in situ or ex situ transesterification; and improve its cost competitiveness as an abundant feedstock source for biofuels production. To reduce production costs and maintain environmental sustainability, influent wastewater and waste lignocellulose biomass hydrolyzate were used as cultivation media and substrate, respectively. However, lignocellulose hydrolyzates also contain degradation by-products such as furfural and acetic acid that are known to exert inhibitory effects on microorganisms; hence their effects on the fermentative performance of activated sludge were investigated and fermentation strategies were proposed and evaluated to counteract the microbial toxicity of these compounds. The utilization rate and efficiency of xylose by activated sludge microorganisms for lipid production was Template Created By: Damen Peterson 2009 also evaluated as pentose sugars such as xylose usually constitute a major percentage of lignocellulose hydrolyzates. Furthermore, variations in the population profile of activated sludge microbiota were determined via 16S rRNA sequence analysis to determine the effect of sugar fermentation at different initial conditions. Results show that activated sludge lipid contents and biodiesel yield could be enhanced by fermentation of sugars at a high initial C:N ratio (70:1). Furfural was found to be highly inhibitory to microbial growth and lipid accumulation while high initial acetic acid concentrations enhanced biomass production but not lipid formation. Xylose was also utilized more efficiently than glucose by the activated sludge microorganisms for biomass and lipid production albeit at relatively slower rates; hence sugar mixtures derived from lignocellulose could be utilized for the process. Semicontinuous and continuous fermentation modes were proposed and evaluated as strategies to reduce inhibitory effect of furfural and acetic acid and improve lipid productivity, but the lack of nutrient supplementation prevented the cultures from sustaining microbial growth and lipid production, leading to cell death and washout. Finally, the reduction in the diversity of the activated sludge microbiota could point to specific microbial strains that are mainly responsible for lipid accumulation. Template Created By: Damen Peterson 2009 DEDICATION This work is dedicated to my parents, who have sacrificed so much that I may have this opportunity to succeed in my all endeavors; to Sandy, my better half and my inspiration; and to God, with whom all things are possible. ii Template Created By: Damen Peterson 2009 ACKNOWLEDGEMENTS The author would like to thank the following agencies for providing the funding for this research: the U.S. Department of Energy, U.S. Environmental Protection Agency, and the National Science Foundation; as well as to the City of Tuscaloosa, AL Hillard Fletcher Wastewater Treatment Plant for providing the activated sludges used in this study. Special thanks are also given to the members of the author’s dissertation committee. First and foremost, to Dr. Rafael Hernandez; a great mentor, teacher, and leader in whom the author looks up to and hopes to model his future academic career after. Many thanks also to Dr. Todd French, who has been a source of invaluable knowledge in the field of microbiology and also in life. Furthermore, the author thanks Dr. Bill Elmore, Dr. Mark Bricka, Dr. Keisha B. Walters, and Dr. Benjamin Magbanua, for all the invaluable inputs, comments, suggestions, and criticisms that contributed greatly for the improvement of this work. This work would not have been possible without the hard work and dedication of the Renewable Fuels and Chemicals Laboratory (RFCL) staff and students who have worked on the project over the four and a half years since its inception. Many thanks to Ms. Linda McFarland, an excellent lab manager who always worked hard to instill in us a sense of discipline in research; Mr. William Holmes, whose expertise in analytical iii Template Created By: Damen Peterson 2009 instrumentation has contributed greatly to the analytical results in this study; the late Dr. Earl Alley, whose extensive knowledge in analytical chemistry continues to inspire and influence the group’s research in that aspect and everything else; and to the following undergraduate and high school students: Colin Gurtowski, Marlene Trillo, Courtney Ramsey, Otis Malone, Vicdaly Williams, Leni Alex, Monica Haque, and David Batchelor. Many thanks are also given to the rest of the RFCL staff and students, current and former, for all the support and likewise the invaluable comments, criticisms, and suggestions that have contributed in one way or another to the development of this work: Dr. Darrell Sparks (now with the Department of Biochemistry and Molecular Biology, Mississippi State University), Dr. Patrisha Pham, Mr. Jimmie Cain, Dr. Alexei Iretski (Lake Superior State University, Sault Ste. Marie, MI), Jacqueline Hall, Mary Hetrick, Maria Paraschivescu, Emmanuel Revellame, Adebola Coker, and Scott Crymble. The author also thanks the U.S. Environmental Protection Agency, National Risk Management Research Laboratory (NRMRL) group in Cincinnati, OH composed of Dr. Jorge W. Santo Domingo, Dr. Mark Meckes, Dr. Hodon Ryu, and Brandon Iker for performing most of the microbial community analysis and for their hospitality during the author’s stay in Cincinnati to undergo training on molecular methods. Special thanks also to Jacqueline Hall and Mary Hetrick, who have likewise contributed greatly to this part of the overall work and for being such great company during the 10-day training period at the U.S. EPA Cincinnati office. The author would also like to thank the faculty and staff of the Dave C. Swalm School of Chemical Engineering, most especially to Ms. Sandra Shumaker and Ms. Ellen Weeks for all the purchase and trip requests that they made and for their extreme iv Template Created By: Damen Peterson 2009 patience; to Mr. Mike Hillhouse, Ms. Sherre Denson, and Dr. Bill Elmore, Interim Director of the Department. Special thanks are given to Dr. Chay Binh Pham and Dr Laura Pham, for all their support and for being instrumental in