EVALUATION OF BIOLOGICAL STRATEGIES TO ENHANCE HYDROLYSIS DURING ANAEROBIC DIGESTION OF COMPLEX WASTE

Abstract: Anaerobic digestion (AD) offers considerable potential in the production of energy from renewable sources, and is being increasingly recognised as a sustainable technology for the treatment of waste. The production of energy from waste such as municipal solid waste (MSW), lipid-rich waste, and agricultural biomass and waste may play an important role in the production of renewable energy given the huge amounts of these types of waste generated annually. Since it is a biological process, the overall efficiency of the AD process depends on the efficiency of the interactions between the different microorganisms. The first step in AD consists of hydrolysis and solubilization of the particulate substrate. This step may be limiting in a number of situations causing a bottleneck in the overall process. One of the aims of the work presented in this thesis was to improve the understanding of the factors limiting the hydrolytic step and their influence on the overall process when dealing with complex substrates containing lipids and lignocellulose. The significance of the different factors is discussed. The hydrolysis of lipids was found to be limiting under the conditions studied. The mechanism of inhibition of methane production seemed to be the same for all the concentrations of lipid investigated, and inhibition was primarily due to transport limitation phenomena. The major obstacle to methane production was found to be the long-chain fatty acids (LCFAs) formed. The digestion process was not permanently inhibited, even for concentrations of 6500 mg/l, and the percentage of biomethanation was above 90% for all concentrations of lipids. Hydrolysis was also found to be limiting regarding lignocellulose, mainly due to enzyme accessibility problems associated with the composition and structure of lignocellulosic substrates. An increase in the abundance of the cellulolytic microorganisms belonging to the genus Clostridium, was observed when the hydrolysis rate became limiting for the overall degradation process, confirming that hydrolysis was limiting. Another aim of this work was to develop and evaluate biological strategies to overcome the limitations described above. Enzyme addition was evaluated for lignocellulosic substrates, while enzyme addition and bioaugmentation were investigated for lipid-rich substrates. For the lignocellulosic substrate, the addition of cellulases and ?-glucosidase to the residual lignocellulosic fraction of source-sorted MSW, after digestion of the easily degradable organic fraction in a two-stage process, resulted in a 34% increase in the leached soluble organic compounds indicating that this treatment strategy is promising. Bioaugmentation of a one-stage digester with Clostridium lundense, a new anaerobic lipolytic species isolated from bovine rumen, resulted in a higher methane production rate and in a reduction in the digestion time. However, the overall advantages of this strategy may be reduced depending on the composition of the substrate due to the presence of the LCFAs. Finally, the application of fluorescence in situ hybridization (FISH) as a tool for monitoring the microorganisms involved in the AD process was evaluated. FISH allowed a relation to be established between the microbial community and process performance in the hydrolytic stage of two-stage digestion.