Development of efficient tools for monitoring and improvement of biogas production

Abstract

Biomass from energy crops and organic waste is one of the most important renewable energy sources and can be used for the production of biogas. Biogas formation is based on the fermentation of organic matter and is performed in biogas plants that generate electric power and heat. Many groups of microorganisms are involved in the anaerobic degradation of organic material and in the formation of biogas, which proceeds in the four interdependent steps of hydrolysis, acidogenesis, acetogenesis and methanogenesis. Thus, the synthesis of biogas depends on a variety of microorganisms and includes a huge number of biochemical reactions. However, the weakest link of the microbial anaerobic degradation chain determines the performance and the speed of the overall system. This bottle neck has to be determined to specifically optimize the biogas production process. <br /> So far, it is only possible to analyze the overall process performance by analytical monitoring parameters, e.g. biogas formation, VFA concentration, pH, and buffer capacity. Nevertheless, the biochemical bottlenecks are not identified yet and a differential analysis of metabolic activities of the microorganism involved in biogas formation is not possible. Therefore, the aims of this thesis focused on the development of monitoring strategies for the quantification of metabolic capacities of the microorganisms to identify biochemical bottlenecks and to improve the efficiency of biogas production. In the <strong>first chapter</strong> an enzymatic test system is presented for the quantification of methanogenic archaea, the most important microbial group in biogas formation. The analysis of their metabolic activity was based on the heterodisulfide reductase (Hdr), a key enzyme in all methanogenic pathways. Using a rapid enzymatic test system, the activity of the Hdr was detected in cell free extract prepared from biogas sludge which enabled the specific quantification of all methanogenic archaea involved in the anaerobic degradation process. In the second test system the different localization of the Hdr within the methanogenic cells allowed to quantify the metabolic activity of both groups of methanogenic organisms. It became evident that one third of the total Hdr activity was found in the membrane fraction representing aceticlastic methanogens. The cytoplasmic fraction contained two third of the total Hdr activity that derived from hydrogenotrophic methanogens. <br /><strong>Chapter 2</strong> comprises the results on the analysis of the metabolic activity of all microbial degradation steps in biogas sludge to shed light on the question which group of organism constitutes the bottleneck in the anaerobic breakdown of organic material. Biogas sludge was incubated and analyzed in anaerobic small-scale batch reactors within 24 h and the process was analyzed by analytical parameters. CH₄ production was examined after supplementation of biogas sludge with substrates (butyrate, propionate, ethanol, acetate, H₂+CO₂) for the microorganisms involved in the different digestion levels. A significant increase of CH₄ formation was measured when sludge from different biogas plants was supplemented with acetate or ethanol. These results led to the conclusion that aceticlastic methanogenesis and syntrophic ethanol-oxidation did not constitute the biochemical bottleneck during biogas formation, respectively. <br /> Increasing CH₄ formation caused by ethanol addition was also analyzed in small-scale continuous reactors filled with biogas sludge to determine the influence of ethanol on CH₄ production over longer time periods (<strong>Chapter 3</strong>). Pulsed ethanol supplementation and continuous addition of ethanol or ethanolic solutions (e.g. beer) to the reactors led to significantly increased biomethanation. The biogas formation increased directly after the addition of ethanol or ethanolic solutions. Thus, an adjustment of CH₄ production to fluctuant power demands is possible to ensure power supply in times of daily or seasonal peak loads.<br /> The study presented in <strong>Chapter 4</strong> allows to quantify the metabolic performance of microorganisms involved in different digestion levels in biogas plants and enables to identify the weakest link of the microbial anaerobic degradation chain. A test system referred to as <u>BEAP</u> profile was developed which is based on the addition of intermediate substrates of prokaryotes involved in the different digestion levels. Supplementation of biogas sludge samples with butyrate (<u>B</u>CON), ethanol (<u>E</u>CON), acetate (<u>A</u>CON) or propionate (<u>P</u>CON) and subsequently the analysis of CH₄ formation in comparison to control samples without supplementation enabled to characterize the performance of the degradation process by rapid testing of metabolic activities of the microorganisms involved in biogas formation. Furthermore, a differentiation between specific BEAP profiles was possible for standard biogas plants and for biogas reactors with process incidents (beginning of NH₄+-N intoxication, start of acidification, insufficient hydrolysis and potential mycotoxin effects). Moreover, at the beginning of NH₄+-N intoxication the BEAP profiles function as an early warning system to predict critical NH₄+-N concentration thresholds leading to a drop of CH₄ formation in commercial agricultural biogas plants.