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Outdoor Cultivation of Chlorella Vulgaris - Assessment of Yield Influencing Parameters and Application As Biogas Substrate

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Outdoor Cultivation of Chlorella Vulgaris - Assessment of Yield Influencing Parameters and Application As Biogas Substrate Outdoor cultivation of Chlorella vulgaris - Assessment of yield influencing parameters and application as biogas substrate Vom Promotionsausschuss der Technischen Universität Hamburg-Harburg zur Erlangung des akademischen Grades Doktorin der Naturwissenschaften (Dr. rer. nat.) genehmigte Dissertation von Stephanie Brandt aus Helmstedt 2015 Gutachter Prof. Dr.-Ing. Martin Kaltschmitt Prof. Dr. rer. nat. Andreas Liese Tag der mündlichen Prüfung 06. Januar 2015 Danksagung Mein besonderer Dank gilt Herrn Prof. Dr.-Ing. Martin Kaltschmitt für die wissenschaftliche Betreuung meiner Arbeit, für wertvolle Anregungen und Diskussionen und für die volle Unter- stützung. Herrn Prof. Dr. rer. nat. Andreas Liese danke ich für seine Bereitschaft als Korreferent meine Arbeit zu begutachten. Weiterhin gilt mein Dank Frau Prof. Dr.-Ing. Kerstin Kuchta, die mich in ihrer Arbeitsgruppe aufgenommen und stets bei meiner Arbeit unterstützt hat. Ein herzliches Dankeschön geht auch an all die anderen Kolleginnen und Kollegen aus der Schloßstraße, insbesondere Jörn Heeren- klage, für die Hilfestellungen im Labor und bei den Versuchen und für erbauliche Flurdiskussio- nen und kurzweilige Kaffeepausen. Frau Dr. Barbara Klippel danke ich für die Unterstützung bei den Enzymversuchen. Ohne meine Kollegen und Kolleginnen der E.ON Hanse AG wäre diese Doktorarbeit nicht zu- stande gekommen. Insbesondere gilt mein Dank Ralf, Claus, Thomas, Kai-Uwe, Nebo, Jasmina, Uwe, Ahmet, Kai, Roman, Arno, Herrn Behrens, Mario und Melanie. Das waren ein paar wirklich gute Jahre mit Euch! Ein weiterer Dank geht an die Arbeitsgruppe von Frau Dr. Ulrike Schmid-Staiger vom Fraunhofer- Institut für Grenzflächen- und Bioverfahrenstechnik IGB in Stuttgart für die Ermöglichung der Fettsäureanalytik und für interessante fachliche Gespräche (Danke auch Dir, Ronja!). Herrn Prof. Dr. Walter Trösch, Herrn Dr. Peter Ripplinger und den Mitarbeitern der Subitec GmbH danke ich für die wertvollen Diskussionen zur Kultivierung von Mikroalgen in FPA-Reaktoren und für die gute Zusammenarbeit. Anja Schwenzfeier danke ich für die Versuche zur Extraktion von löslichen Proteinen aus der Algenbiomasse. Dr. Mirko Lunau danke ich besonders für seine Einführung in die Durchflußzy- 3 i tometrie und seine Unterstützung. Unsere Gespräche gaben mir immer neuen Mut. Es hat einfach Spaß gemacht! Mein größter Dank geht an Gunnar, Karla und Jakob, meine Familie. Danke für Erdung, Antrieb, Unterstützung und vor allem für Verständnis während der ganzen Jahre. Ich bin jeden Tag dankbar, dass es Euch gibt! Table of Contents 1 Introduction 2 1.1 Background . .3 1.2 Objectives . .6 1.3 General approach . .7 2 Fundamentals 9 2.1 Microalgae . .9 2.1.1 Chlorella vulgaris ............................9 2.1.2 Growth characteristics and requirements . 13 2.1.3 Products . 15 2.2 Cultivation systems . 19 2.2.1 Process technology . 19 2.2.2 Operation modes . 23 3 Material and Methods 24 3.1 Organism, culture conditions, and inoculum . 24 3.2 The outdoor pilot plant . 25 3.2.1 The 180 L Flat-Panel Airlift photobioreactor . 25 ii Table of Contents iii 3.2.2 Design of the pilot plant . 27 3.2.3 Operation of the pilot plant . 27 3.2.4 Sources of error . 30 3.3 Analytical and statistical methods . 31 3.3.1 Biotic parameters . 31 3.3.2 Abiotic parameters . 38 3.3.3 Statistical methods . 43 3.4 Experimental designs and procedures . 44 3.4.1 Experiment 1: Effect of light and temperature on productivity and com- position . 45 3.4.2 Experiment 2: Nitrate and phosphate uptake and the fate of carbon . 46 3.4.3 Experiment 3: Flue gas as a carbon and nitrogen source . 46 3.4.4 Experiment 4: Microalgae as a feedstock for anaerobic digestion . 49 4 Results and Discussion 53 4.1 Experiment 1: Effect of light and temperature on productivity and composition 53 4.1.1 Effect of varying light intensity and temperature on the temporal dynam- ics of algal growth and composition . 53 4.1.1.1 Results . 53 4.1.1.2 Discussion . 56 4.1.2 Impacts of environmental conditions on growth . 61 4.1.2.1 Results . 61 4.1.2.2 Discussion . 64 4.2 Experiment 2: Nitrate and phosphate uptake and the fate of carbon . 68 4.2.1 Uptake and assimilation of nitrate and phosphate . 68 4.2.1.1 Results . 68 4.2.1.2 Discussion . 69 4.2.2 Fate of dissolved organic carbon . 70 4.2.2.1 Results . 70 4.2.2.2 Discussion . 71 4.3 Experiment 3: Flue gas as a carbon and nitrogen source . 75 4.3.1 Proving the system comparability . 75 4.3.1.1 Results . 76 4.3.1.2 Discussion . 79 4.3.2 Effect of flue gas on the productivity and composition . 79 4.3.2.1 Results . 80 4.3.2.2 Discussion . 84 4.3.3 Removal of NOx and CO2 from flue gas . 86 4.3.3.1 Results . 86 4.3.3.2 Discussion . 88 4.4 Experiment 4: Microalgae as a feedstock for anaerobic digestion . 89 4.4.1 Prerequisites for anaerobic digestion experiments . 90 4.4.1.1 Results . 90 4.4.1.2 Discussion . 91 4.4.2 Digestion of pre-treated biomass . 92 4.4.2.1 Results . 92 4.4.2.2 Discussion . 96 4.4.3 Digestion of residual microalgae biomass . 101 4.4.3.1 Results . 101 4.4.3.2 Discussion . 102 5 Summary and Outlook 104 List of Abbreviations Abbreviation Full name A Autotrophic growth ALA α-Linolenic acid ANOVA Analysis of variance ATP Adenosine triphosphate cs Closed system Cel Cellulase Cel/Pec Cellulase/Pectinase CHP Combined heat and power DHA Docosahexanoic acid DMSO Dimethyl sulfoxide DNA Deoxyribonucleic acid DTT Dithiothreitol DWD Deutscher Wetterdienst EDTA Ethylenediaminetetraacetic acid F Ratio of mean squares (ANOVA) FAME Fatty acid methyl ester FPA Flat-Plate Airlift GC Gas chromatography GF Glass fiber GHG Greenhouse gas GMP Good manufacturing practice H Heterotrophic growth M Mixotrophic growth v vi LIST OF ABBREVIATIONS Abbreviation Full name Max Maximum Min Minimum N-Prot Nitrogen-Protein os Open system ! Omega p Probability PBR Photobioreactor PCA Principal component analysis PC1 Principal component 1 PC2 Principal component 2 PC1ro Principal component 1 after applying a varimax rotation PC2ro Principal component 2 after applying a varimax rotation Pec Pectinase PUFA Polyunsaturated fatty acid r Correlation coefficient RNA Ribonucleic acid SAG Culture Collection of Algae at Göttingen University Std Standard deviation STP Standard temperature and pressure TAGs Triacylglycerols Tris Tris(hydroxymethyl)aminomethane TUHH Hamburg University of Technology VDI Verein Deutscher Ingenieure WTW Wissenschaftlich-Technische Werkstätten GmbH Parameter Full name α alpha (Parameter in equation 3.2) B Biomass concentration CellCAlg Cell counts (microalgae) CellCBac Cell counts (bacteria) Cl Cloudiness CO2;in Carbon dioxide content in the flue gas stream (inlet) CO2;out Carbon dioxide content in the flue gas stream (outlet) COD Chemical oxygen demand DegreeCO2 Degree of decarbonisation DIN Dissolved inorganic nitrogen DIP Dissolved inorganic phosphorus LIST OF ABBREVIATIONS vii Parameter Full name DLIver Vertically measured daily light intensity DOC Dissolved organic carbon FA Fatty acid FCO2 Carbon dioxide biofixation rate F racBac Total bacterial fraction LA Light availability lBac Length of a bacterial cell LIver Vertically measured light intensity MaxL Vertically measured maximum light intensity MaxT Maximum temperature MC Molar mass of carbon MCO2 Molar mass of carbon dioxide MN Molar mass of nitrogen MNO3 Molar mass of nitrate MP Molar mass of phosphorus MPO4 Molar mass of phosphate MeanT Mean temperature MinT Minimum temperature OD Optical density OD750 Optical density at a wavelength of 750 nm OP D Optimal population density OP P Optimal population productivity OC Organic carbon OLA Optimal light availability ON Organic nitrogen P Productivity P OC Particulate organic carbon P ON Particulate organic nitrogen P OP Particulate organic phosphorus rAlg Radius of a microalgal cell rBac Radius of a bacterial cell sCOD Soluble chemical oxygen demand SD Sun duration SurfReactor Reactor surface area T Temperature T OC Total organic carbon TS Total solids viii LIST OF ABBREVIATIONS Parameter Full name T SVAlg Total specific biovolume (microalgae) T SVBac Total specific biovolume (bacteria) T V ar Temperature variance VAlg Biovolume of a microalgal cell VBac Biovolume of a bacterial cell VReactor Reactor volume VS Volatile solids Units Full name d Day °C Degree Celsius g Gram h Hour Hz Hertz kg Kilogram KJ Kilojoule km2 Square kilometre l Length L Litre m Metre M Mol m2 Square metre m3 Cubic metre µE Microeinstein µg Microgram µL Microlitre µm Micrometre µm3 Cubic micrometre µmol Micromole mg Milligram min Minute mL Millilitre mm Millimeter mM Millimolar MW Megawatt nm Nanometre LIST OF ABBREVIATIONS 1 Units Full name % Per cent ppm Parts per million rpm Rounds per minute t Time x g Times gravity Vol.-% Volume per cent vvm Volume per volume per minute Chapter 1 Introduction The world energy consumption keeps rising since the industrial revolution and is expected to increase further by 53 % between 2008 and 2035 (EIA, 2011). A large part of the increase will be attributed to an increasing population and a fast economic growth of countries like China or India. Although renewable energy is the fastest growing energy source worldwide, it is pre- dicted that fossil fuels will still cover 80 % of the overall energy demand in 2035 (EIA, 2011). However, fossil fuel reserves are limited and the production of oil and gas is becoming increas- ingly demanding and, thus, expensive (Murray and King, 2012; ter Veld, 2012). Securing energy supply will therefore become a major challenge in the future. Closely linked to the depleting resources of fossil fuels is the rising carbon dioxide concentration in the atmosphere, which contributes to anthropogenic global warming (IPCC, 2007; Murray and King, 2012). Currently, many of the renewable energy sources under development focus on the electricity mar- ket (e.g.
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