Research Collection Doctoral Thesis Investigating the mode of action of fatty acids against rumen methanogens Author(s): Zhou, Xuan Publication Date: 2015 Permanent Link: https://doi.org/10.3929/ethz-a-010510909 Rights / License: In Copyright - Non-Commercial Use Permitted This page was generated automatically upon download from the ETH Zurich Research Collection. For more information please consult the Terms of use. ETH Library DISS. ETH NO.22675 Investigating the mode of action of fatty acids against rumen methanogens A dissertation submitted to attain the degree of DOCTOR OF SCIENCES of ETH ZURICH (Dr. Sc. ETH Zürich) Presented by Xuan Zhou Nanjing Agricultural University, China MSc Agr. Born 09.03.1986 Citizen of China Accepted on the recommendation of Prof. Dr. Michael Kreuzer, examiner Prof. Dr. Leo Meile, co-examiner Dr. Angela Schwarm, co-examiner Dr. Johanna Zeitz, co-examiner 2015 As tiny as me In the giant universe Home not to everywhere However the most dangerous and hostile corners As dark as dungeon As radiated as nuclear As acid as king's water As alkaline as Alkali Lake As hot as black smoker As cold as iceberg Where I survive, my families thrive As ancient as me Witness the footprints of history As simple as me Delicate my whole life to dancing on evolutionary stage Catch my tail if you could human Wizard will reveal you a secret On where did you come from On where are you heading to ---A little piece of poem dedicate to Archaea Table of content Table of content ...................................................................................................................................... I List of Abbreviations ........................................................................................................................... IV List of Tables ......................................................................................................................................... V List of Figures ...................................................................................................................................... VI Summary .................................................................................................................................................7 Zusammenfassung ..................................................................................................................................9 1 General introduction ...............................................................................................................11 1.1 Methane emission from ruminants ............................................................................................12 1.2 Rumen methanogens .................................................................................................................12 1.3 Diet supplements with antimethanogenic compounds ..............................................................13 1.3.1 Effect of saturated fatty acids on methanogens .........................................................................14 1.3.2 Characteristics of fatty acids .....................................................................................................16 1.3.3 Effects of fatty acids on other organisms ..................................................................................17 1.3.4 The effects of fatty acids on other rumen protozoa ...................................................................18 1.4 Characteristics of M. ruminantium and its methanogenesis ......................................................18 1.5 Research topic and hypotheses tested in the present doctoral thesis .........................................21 2 The effect of saturated fatty acids on methanogenesis and cell viability of Methanobrevibacter ruminantium ...........................................................................................22 2.1 Abstract .....................................................................................................................................23 2.2 Introduction ...............................................................................................................................23 2.3 Materials and methods ..............................................................................................................25 2.3.1 Strain and growth conditions ....................................................................................................25 2.3.2 Experiment 1 .............................................................................................................................26 2.3.3 Experiments 2 and 3 ..................................................................................................................27 2.3.4 Statistical analysis .....................................................................................................................28 2.4 Results .......................................................................................................................................29 2.4.1 Inhibition of methane production of M. ruminantium by SFA as depending on dose in experiment 1 ..............................................................................................................................29 + 2.4.2 Influence of C 12 on methane production, K leakage and cell viability in experiment 2 ..........30 2.4.3 Influence of SFA on methane production, K + leakage, and cell viability in experiment 3 .......30 2.5 Discussion .................................................................................................................................32 2.5.1 Efficiency of saturated fatty acids to inhibit methanogenesis in Methanobrevibacter ruminantium ..............................................................................................................................32 2.5.2 Indications for modes of action of saturated fatty acids ............................................................33 I 2.6 Conclusion ................................................................................................................................ 34 3 Inhibitory effects of saturated fatty acids on methane production by methanogenic archaea .................................................................................................................................... 35 3.1 Abstract .................................................................................................................................... 36 3.2 Introduction .............................................................................................................................. 36 3.3 Materials and methods .............................................................................................................. 37 3.3.1 Experimental microbes and cultivation .................................................................................... 37 3.3.2 Experiment 1 ............................................................................................................................ 38 3.3.3 Experiment 2 ............................................................................................................................ 38 3.3.4 Laboratory and statistical analysis ............................................................................................ 39 3.4 Results ...................................................................................................................................... 39 3.4.1 Experiment 1 ............................................................................................................................ 39 3.4.2 Experiment 2 ............................................................................................................................ 42 3.5 Discussion ................................................................................................................................ 43 3.5.1 Importance of fatty acid chain length, cultivation temperature and substrate for the anti- methanogenic effect of SFA on the archaea ............................................................................. 43 3.5.2 Potential modes of action regarding the inhibitory effect of the saturated fatty acids ............. 44 3.6 Conclusions .............................................................................................................................. 45 4 Influence of pH on the inhibitory effect of fatty acids on the rumen methanogen Methanobrevibacter ruminantium .......................................................................................... 46 4.1 Abstract .................................................................................................................................... 47 4.2 Introduction .............................................................................................................................. 47 4.3 Materials and methods .............................................................................................................. 49 4.3.1 Strain and growth conditions .................................................................................................... 49 4.3.2 Pretest: Identification of the optimal experimental conditions and of the range of pH values tolerated by M. ruminantium .................................................................................................... 50 4.3.3 Main experiment: Effect of differentially protonated C 12 and C 14 on M. ruminantium ............ 51 4.3.4 Statistical analysis ...................................................................................................................