Kinetics of propionate conversion in anaerobic continuously stirred tank reactors.

The kinetic parameters of anaerobic propionate degradation by biomass from 7 continuously stirred tank reactors differing in temperature, hydraulic retention time and substrate composition were investigated. In substrate-depletion experiments (batch) the maximum propionate degradation rate, Amax, and the half saturation constant, Km, were initially estimated by applying the integrated Michaelis-Menten equation. Amax was in the range from 22.8 to 29.1 micromol gVS(-1) h(-1) while Km was in the range from 0.46-0.95 mM. In general, Amax gave a good reflection of the reactor performances. Secondly, the accuracy of the applied method was evaluated by use of radiotracer methodology. Amax was found to be 14-15% lower in the substrate-depletion experiment than in the radioisotope experiment due to endogenous propionate production. By including the endogenous propionate production, a 42-49% lower Km was estimated. The results demonstrate that the rate of endogenous substrate (propionate) production should be taken into account when estimating kinetic parameters in biomass from manure-based anaerobic reactors.

Thermal pretreatment of the solid fraction of manure: impact on the biogas reactor performance and microbial community.

Application of thermal treatment at 100-140 degrees C as a pretreatment method prior to anaerobic digestion of a mixture of cattle and swine manure was investigated. In a batch test, biogasification of manure with thermally pretreated solid fraction proceeded faster and resulted in the increase of methane yield. The performances of two thermophilic continuously stirred tank reactors (CSTR) treating manure with solid fraction pretreated for 40 minutes at 140 degrees C and non-treated manure were compared. The digester fed with the thermally pretreated manure had a higher methane productivity and an improved removal of the volatile solids (VS). The properties of microbial communities of both reactors were analysed. The specific methanogenic activity (SMA) test showed that both biomasses had significant activity towards hydrogen and formate, while the activity with the VFA - acetate, propionate and butyrate - was low. The kinetic parameters of the VFA conversion revealed a reduced affinity of the microbial community from the CSTR fed with thermally pre-treated manure for acetate, propionate and butyrate. The bacterial and archaeal populations identified by t-RLFP analysis of 16S rRNA genes were found to be identical in both systems. However, a change in the abundance of the species present was detected.

Comparison of two-stage thermophilic (68 degrees C/55 degrees C) anaerobic digestion with one-stage thermophilic (55 degrees C) digestion of cattle manure.

A two-stage 68 degrees C/55 degrees C anaerobic degradation process for treatment of cattle manure was studied. In batch experiments, an increase of the specific methane yield, ranging from 24% to 56%, was obtained when cattle manure and its fractions (fibers and liquid) were pretreated at 68 degrees C for periods of 36, 108, and 168 h, and subsequently digested at 55 degrees C. In a lab-scale experiment, the performance of a two-stage reactor system, consisting of a digester operating at 68 degrees C with a hydraulic retention time (HRT) of 3 days, connected to a 55 degrees C reactor with 12-day HRT, was compared with a conventional single-stage reactor running at 55 degrees C with 15-days HRT. When an organic loading of 3 g volatile solids (VS) per liter per day was applied, the two-stage setup had a 6% to 8% higher specific methane yield and a 9% more effective VS-removal than the conventional single-stage reactor. The 68 degrees C reactor generated 7% to 9% of the total amount of methane of the two-stage system and maintained a volatile fatty acids (VFA) concentration of 4.0 to 4.4 g acetate per liter. Population size and activity of aceticlastic methanogens, syntrophic bacteria, and hydrolytic/fermentative bacteria were significantly lower in the 68 degrees C reactor than in the 55 degrees C reactors. The density levels of methanogens utilizing H2/CO2 or formate were, however, in the same range for all reactors, although the degradation of these substrates was significantly lower in the 68 degrees C reactor than in the 55 degrees C reactors. Temporal temperature gradient electrophoresis profiles (TTGE) of the 68 degrees C reactor demonstrated a stable bacterial community along with a less divergent community of archaeal species.

Anaerobic digestion of manure and mixture of manure with lipids: biogas reactor performance and microbial community analysis.

Anaerobic digestion of cattle manure and a mixture of cattle manure with glycerol trioleate (GTO) was studied in lab-scale, continuously stirred tank reactors (CSTR) operated at 37 degrees C. The reactor codigesting manure and lipids exhibited a significantly higher specific methane yield and a higher removal of VS than the reactor treating manure. Microbial population analysis done by cultivation--most probable number (MPN) test and specific methanogenic activity (SMA) measurement, revealed higher MPN and increased SMA of methanogenic populations of biomass from the reactor codigesting manure and lipids. Spatial microbial distribution and activity was studied in digested materials fractionated into size of particles > 200 microm, 50-200 microm and 0.45-50 microm. With manure, the main pool of methanogenic activity from propionate, butyrate and hydrogen was associated with the particles > 200 microm, while the activity of acetotrophic methanogens was uniformly distributed in all fractions. When digesting manure and lipids, an enhanced methanogenesis was detected both for particles > 200 microm and the 50-200 microm fraction. The molecular methods--temperature gradient gel electrophoresis (TGGE), cloning library and sequencing of 16S rDNA--showed presence of a restricted number of archaeal species in both reactors. The vast majority of clones was phylogenetically most closely related to Methanosarcina siciliae.

State of the art and future perspectives of thermophilic anaerobic digestion.

The state of the art of thermophilic digestion is discussed. Thermophilic digestion is a well established technology in Europe for treatment of mixtures of waste in common large scale biogas plants or for treatment of the organic fraction of municipal solid waste. Due to a large number of failures over time with thermophilic digestion of sewage sludge this process has lost its appeal in the USA. New demands on sanitation of biosolids before land use will, however, bring the attention back to the use of elevated temperatures during sludge stabilization. In the paper we show how the use of a start-up strategy based on the actual activity of key microbes can be used to ensure proper and fast transfer of mesophilic digesters into thermophilic operation. Extreme thermophilic temperatures of 65 degrees C or more may be necessary in the future to meet the demands for full sanitation of the waste material before final disposal. We show data of anaerobic digestion at extreme thermophilic temperatures.

Effect of temperature increase from 55 to 65 degrees C on performance and microbial population dynamics of an anaerobic reactor treating cattle manure.

The effect of a temperature increase from 55 to 65 degrees C on process performance and microbial population dynamics were investigated in thermophilic, lab-scale, continuously stirred tank reactors. The reactors had a working volume of 31 and were fed with cattle manure at an organic loading rate of 3 g VS/l reactor volume/d. The hydraulic retention time in the reactors was 15 days. A stable reactor performance was obtained for periods of three retention times both at 55 degrees C and 65 degrees C. At 65 degrees C methane yield stabilized at approximately 165ml/g VS/d compared to 200 ml/g VS/d at 55 degrees C. Simultaneously, the level of total volatile fatty acids, VFA, increased from being below 0.3 g/l to 1.8-2.4 g acetate/l. The specific methanogenic activities (SMA) of biomass from the reactors were measured with acetate, propionate, butyrate, hydrogen, formate and glucose. At 65 degrees C. a decreased activity was found for glucose-, acetate-, butyrate- and formate-utilizers and no significant activity was measured with propionate. Only the hydrogen-consuming methanogens showed an enhanced activity at 65 degrees C. Numbers of cultivable methanogens, estimated by the most probable number (MPN) method, were significantly lower on glucose, acetate and butyrate at the increased operational temperature, while the numbers of hydrogenotrophic methanogens remained unchanged. No viable propionate-degrading bacteria were enriched at 65 C. Use of ribosomal oligonucleotide probes showed that an increase in temperature resulted in a decreased contribution of the rRNA of the domain bacteria from 74-79 to 57-62% of the universal probe, while the rRNA of the domain archaea, increased from 18-23 to 34-36%.

Growth kinetics of thermophilic Methanosarcina spp. isolated from full-scale biogas plants treating animal manures.

This study determines the growth kinetics of thermophilic strains of Methanosarcina spp. from full-scale thermophilic biogas plants. The complete set of kinetic parameters, including maximum specific growth rate µ(max), half saturation constant K(S), acetate threshold concentration and cell growth yield Y(X/S), were determined for six Methanosarcina strains newly isolated from full-scale reactors and the type strain Methanosarcina thermophila TM-1(T). The kinetic experiments were performed in media supplemented with acetate and activated carbon at the optimum growth temperatures of the individual strains, 50-55 degrees C. The µ(max) values of the isolates were in the range of 0.044-0.064 h(-1), the K(S) ranged from 6.5 to 24.7 mM acetate and the threshold for acetate utilization from 0.11 to 0.40 mM. The cell growth yields of the strains were between 0.78 and 2.97 g dry weight cells mol(-1) acetate. The six isolates exhibited significantly higher µ(max) and had higher affinity to acetate than the type strain M. thermophila TM-1(T). Generally, the affinities of thermophilic Methanosarcina strains tested in this study cover a similar range to those reported in the literature for mesophilic Methanosarcina spp. with acetate as substrate. The strains isolated from plants treating mixtures of animal manures and industrial organic wastes had higher affinity for acetate and lower thresholds than strains isolated from reactors operating solely on manures.

Acetate conversion in anaerobic biogas reactors: traditional and molecular tools for studying this important group of anaerobic microorganisms.

Different methods were applied to study the role of aceticlastic methanogens in biogas reactors treating solid waste and wastewater. We used traditional microbiological methods, immunological and 16S rRNA ribosomal probes for detection of the methanogens. Using this approach we identified the methanogenic spp. and their activity. In biofilm systems, such as the UASB reactors the presence of the two aceticlastic methanogens could be correlated to the difference in the kinetic properties of the two species. In biogas reactors treating solid wastes, such as manure or mixture of manure and organic industrial waste, only Methanosarcina spp. were identified. Methanosarcina spp. isolated from different plants had different kinetics depending on their origin. Relating the reactor performance data to measurement of the activity by conventional microbiological methods gave a good indication of the microbial status of specific trophic groups. 16S rRNA probing confirmed these observations and gave a more detailed picture of the microbial groups present.