Anaerobic digestion of the microalga Spirulina at alkaline conditions (pH~ 10; 2.0 M Na+) Biogas ...

219 Pages·2015·6.22 MB·English

by  Vimac Nolla

Anaerobic digestion of the microalga Spirulina
+
at alkaline conditions (pH~10; 2.0 M Na )
Biogas production and metagenome analysis

Dissertation

In accordance with the requirements for the academic degree of
Doctor of Natural Sciences (Dr. rer. nat.)
Submitted by

Vímac Nolla Ardèvol

Completed at the Faculty of Biology, Bielefeld University,
under the co-supervision of Prof. Dr. Olaf Kruse,
Chair of Algae Biotechnology & Bioenergy,
and Prof. Dr. Marc Strous,
CAIP Research Chair,
Leader of the Energy Bioengineering Group,
Faculty of Science, University of Calgary
October 2014

Main reviewer: Prof. Dr. Olaf Kruse (Universität Bielefeld)
Secondary reviewer: Prof. Dr. Mark van Loosdrecht (Delft University)

This Doctoral Theses has been Printed on “non-ageing paper °° ISO 9706”

Para mi familia y amigos
To my friends and family

Acknowledgments

This thesis would have not been possible without my supervisor, Prof. Dr. Marc
Strous, who gave me the opportunity to work on a fascinating and challenging
project. Thank you for believing in me and for all the helpful advice that you have
given me throughout this three years. My gratitude goes also to Prof. Dr. Olaf Kruse
who kindly “adopted” me during the last months of my thesis. I also have to deeply
thank Dr. Halina Tegetmeyer for helping me during this journey. Thank you for being
so patient with all my constant doubts and problems with the bioinformatic analysis
and for reading and re-reading all my manuscripts, reports, and posters. Thank you
also for taking care of the reactors whenever I wasn’t here and for giving me useful
tips and ideas. Without your help this would have been a nightmare. Special mention
for Dr. Dimitry Y Sorokin who kindly provided the soda lake inoculum that made
possible this project.
Thanks a lot to Ines also for taking care of the multiple reactors and for patiently walk
me through all the sequencing protocols. I also have to mention Regina, our
bioinformatician, who wrote most of the scripts that I’ve used and who was always
there whenever a command was not working. You both made my life here much
easier. Many thanks to Viktor Klassen who helped me to solve a few problems with
the reactors and who introduced me to the world of the batch test. Thanks also to
Philipp, Dominik and their master students from the Fermentationstechnik who let me
“occupy” their Gas Chromatographer. I would also like to thank all the members of
the Algae Biotechnology and Bioenergy group for “adopting” me as one more of the
group and inviting me to all the cakes and get together.
Special thanks to Kyle Lauersen with whom I’ve spent most of my time during this
last years and with whom I shared multiple anecdotes and confidences. We will
never forget Russia. Many thanks to Rabeaa, Anh, Mari, Fernando and the rest of
the CLIB-GC PhD students; it has been a pleasure to share this experience with all of
you. I also have to thank the CLIB 2021 Graduate Cluster for funding and for letting
me be part of this extraordinary PhD program. In relation to this, I have to thank Dr.
Iris Brune for making all the CLIB related paperwork much easier and for kindly
helping me whenever I needed some official document. Thanks for making things
much easier.
També he d’agrair al Dr. Sergi Astals i a la Miriam Peces per ajudar-me des de la
distancia, ja fos des de Barcelona o be des de Queensland, amb tots els dubtes que
he tingut i gracies també per ser els meus proveïdors “oficials” de papers
impossibles. Aquesta experiència no hages estat possible sense en Josep Turet,
l’Anna, l’Albert i l’Enric del SART de la Universitat de Vic qui em van introduir al mon
de la digestió anaeròbia i de la biotecnologia ambiental. Moltes gracies de tot cor.
I also have to thank all my friends around the world. Muchas gracias a Illa, Valen,
Pablo, Meeta, Jivi y el resto de los chicos de Barcelona, a Giorgia, Habiba, Petró,
Pratha y demás amigos que tengo perdidos por ahí, sin vuestros sofás, vuestros
ánimos, vuestros skypes y vuestros apoyos durante este tiempo no lo hubiera
conseguido. Gracias de todo corazón
Y por último, no hay palabras suficientes para agradecer el apoyo recibido de Suvi,
Sarjanet, y Amandita. Ha sido un largo viaje de 10 años desde Vic hasta Bielefeld.
Muchas, muchas, muchas gracias por apoyarme en todo. Os quiero.
Un especial recuerdo para mi padre, que esté donde esté seguro que estará
orgulloso.

Thank you all. We made it!!
Gracias a todos. Lo hemos conseguido!!
Table of contents
Summary i
Abbreviations iii
1. Introduction 1
1.1. The anaerobic digestion process 1
1.1.1. Main parameters affecting the anaerobic digestion process 2
1.1.2. Adding value through anaerobic digestion 4
1.2. Microalgae as substrate for anaerobic digestion 5
1.3. Extreme alkaline (soda) lakes 8
1.3.1. Characteristics of soda lakes 8
1.3.2. Soda lakes of the Kulunda steppe (Russia) 9
1.3.3. Microbial diversity of soda lakes 10
1.3.4. Biotechnological applications of haloalkaliphiles 12
1.4. Microbial community of the anaerobic digestion process 13
1.4.1. Microbiology of the anaerobic digestion process 13
1.4.2. Metagenomics to study the anaerobic digestion process 17
2. Challenges 19
3. Objectives 21
4. General Discussion 23
4.1. Anaerobic digestion of Spirulina at mesophilic pH [3] 25
4.2. Selection of alkaline inoculum and substrate [1] 26
4.3. Anaerobic digestion of Spirulina at alkaline conditions (pH~10; 2.0 M Na+) [2, 4, 5] 28
4.3.1. Determination of the optimal hydraulic retention time [2] 29
4.3.2. Determination of the optimal organic loading rate [2] 31
4.3.3. Biogas production under the optimal process parameters [4] 33
4.3.4. Alkaline digestion of other substrates [5] 36
4.3.5. Biogas rich in methane [2,4,5] 38
4.4. Metagenomics of the anaerobic digestion process [3,6] 39
4.4.1. Characteristic taxonomic structure of anaerobic digester microbial
40
communities [3,6]
4.4.2. Identification of specific functional characteristics in the alkaline
42
metagenome [6]
4.4.3. Comparison of metagenomes derived from anaerobic digesters fed with
43
either protein rich or cellulose rich substrates [3]
4.4.4. Can metagenomics facilitate the optimization of the anaerobic digestion
46
process? [3,6]
4.5. Conclusions 48
5. Perspectives 49
6. Publications 51
[1] Activity and diversity of haloalkaliphilic methanogens in Central Asian soda
53
lakes.
[2] Anaerobic digestion of the microalga Spirulina at alkaline conditions (pH~10;
71
2.0 M Na+).
[3] Metagenome analysis and biogas production from the anaerobic digestion of
89
the protein rich microalga Spirulina
7. Unpublished results 119
[4] Anaerobic digestion of Spirulina at alkaline conditions (pH ~10; 2.0 M Na+)
121
from fresh soda lake sediments
[5] Biomethane potential of different substrates at alkaline conditions. 133
[6] Metagenome analysis of the microbial population from an alkaline anaerobic
147
digester fed with the microalga Spirulina.
8. References 177
9. Appendix 189
10. Curriculum Vitae 203