Tailoring wet explosion process parameters for the pretreatment of cocksfoot grass for high sugar yields.

The pretreatment of lignocellulosic biomass is crucial for efficient subsequent enzymatic hydrolysis and ethanol fermentation. In this study, wet explosion (WEx) pretreatment was applied to cocksfoot grass and pretreatment conditions were tailored for maximizing the sugar yields using response surface methodology. The WEx process parameters studied were temperature (160-210 °C), retention time (5-20 min), and dilute sulfuric acid concentration (0.2-0.5 %). The pretreatment parameter set E, applying 210 °C for 5 min and 0.5 % dilute sulfuric acid, was found most suitable for achieving a high glucose release with low formation of by-products. Under these conditions, the cellulose and hemicellulose sugar recovery was 94 % and 70 %, respectively. The efficiency of the enzymatic hydrolysis of cellulose under these conditions was 91 %. On the other hand, the release of pentose sugars was higher when applying less severe pretreatment conditions C (160 °C, 5 min, 0.2 % dilute sulfuric acid). Therefore, the choice of the most suitable pretreatment conditions is depending on the main target product, i.e., hexose or pentose sugars.

Pretreatment as the crucial step for a cellulosic ethanol biorefinery: testing the efficiency of wet explosion on different types of biomass.

The efficiency of wet explosion applied as modified dilute acid pretreatment at previously identified reference conditions (150°C, 0.3% H(2)SO(4), 15 min) was investigated on lucerne, ryegrass, fescue grass, cocksfoot grass, rye fescue, forage grass, and wheat straw in order to identify their potential as feedstock for cellulosic bioethanol production. After pretreatment, cellulose recovery was more than 95% for all biomass while enzymatic convertibility of cellulose ranged from 40% to 80%. Lower enzymatic conversion of cellulose was correlated with higher lignin content of the biomass. Hemicellulose recovery was 81-91% with a final pentose yield of 65-85%. Cocksfoot grass and wheat straw had the highest bioethanol potential of 292 and 308 L/ton DM, respectively. Overall efficiencies were higher than 68% for cocksfoot grass harvested in August, fescue grass, wheat straw, and forage grass while efficiencies were lower than 61% for the other tested biomass resources, making further adjustment of the process parameters necessary.

Microbial community structure and performance of an anaerobic reactor digesting cassava pulp and pig manure.

Microbial community dynamics in response to changes in substrate types (i.e. pig manure (PM), cassava pulp (CP) and mixtures of PM and CP) were investigated in an anaerobic continuously stirred tank reactor (CSTR). Molecular identification of bacterial and archaeal domains were performed, using a 16S rDNA clone library with polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE) screening and phylogenetic analysis. Analysis of bacterial clone libraries revealed that the differences in the community structure corresponded to the substrate types. However, the Bacteroidetes were the most abundant group in all substrates, followed by the Clostridia. With pure PM, the dominant bacterial groups were Bacteroidales, Clostridia and Paludibacter. With a co-substrate, at CP to PM (CP:PM) ratio of 50:50, the sequences analysis revealed the greatest diversity of bacterial communities at class level, and the sequences affiliated with Cytophaga sp. became an exclusive predominant. With CP alone, Bacteroides sp. was the dominant species and this reactor had the lowest diversity of bacteria. Archaea observed in the CSTR fed with all substrate types were Methanosaeta sp., Methanosaeta concilii and Methanospirillum hungatei. Among the Archaea, Methanosaeta sp. was the exclusive predominant. The relative distribution of Archaea also changed regarding to the substrate types.

Improving biogas yields using an innovative concept for conversion of the fiber fraction of manure.

The potential of a new concept to enable economically feasible operation of manure-based biogas plants was investigated at laboratory scale. Wet explosion (WEx) was applied to the residual manure fibers separated after the anaerobic digestion process for enhancing the biogas yield before reintroducing the fiber fraction into the biogas reactor. The increase in methane yield of the digested manure fibers was investigated by applying the WEx treatment under five different process conditions. The WEx treatment at 180 °C and a treatment time of 10 min without addition of oxygen was found to be optimal, resulting in 136% increase in methane yield compared with the untreated digested manure fibers in batch experiments. In a continuous mesophilic reactor process the addition of WEx-treated digested fibers in co-digestion with filtered manure did not show any signs of process inhibition, and the overall methane yield was on average 75% higher than in a control reactor with addition of non-treated digested fibers.

Anaerobic digestion as final step of a cellulosic ethanol biorefinery: Biogas production from fermentation effluent in a UASB reactor-pilot-scale results.

In order to lower the costs for second generation bioethanol from lignocellulosic biomass anaerobic digestion of the effluent from ethanol fermentation was implemented using an upflow anaerobic sludge blanket (UASB) reactor system in a pilot-scale biorefinery plant. Both thermophilic (53 degrees C) and mesophilic (38 degrees C) operation of the UASB reactor was investigated. At an OLR of 3.5 kg-VS/(m(3) day) a methane yield of 340 L/kg-VS was achieved for thermophilic operation (53 degrees C) while 270 L/kg-VS was obtained under mesophilic conditions (38 degrees C). For loading rates higher than 5 kg-VS/(m(3) day) the methane yields were, however, higher under mesophilic conditions compared to thermophilic conditions. The conversion of dissolved organic matter (VS(diss)) was between 68% and 91%. The effluent from the ethanol fermentation showed no signs of toxicity to the anaerobic microorganisms. However, a high content of suspended matter reduced the degradation efficiency. The retention time of the anaerobic system could be reduced from 70 to 7 h by additional removal of suspended matter by clarification. Implementation of the biogas production from the fermentation effluent accounted for about 30% higher carbon utilization in the biorefinery compared to a system with only bioethanol production.

Wet explosion of wheat straw and codigestion with swine manure: effect on the methane productivity.

The continuously increasing demand for renewable energy sources renders anaerobic digestion to one of the most promising technologies for renewable energy production. Twenty-two (22) large-scale biogas plants are currently under operation in Denmark. Most of these plants use manure as the primary feedstock but their economical profitable operation relies on the addition of other biomass products with a high biogas yield. Wheat straw is the major crop residue in Europe and the second largest agricultural residue in the world. So far it has been used in several applications, i.e. pulp and paper making, production of regenerated cellulose fibers as an alternative to wood for cellulose-based materials and ethanol production. The advantage of exploiting wheat straw for various applications is that it is available in considerable quantity and at low-cost. In the present study, the codigestion of swine manure with wheat straw in a continuous operated system was investigated, as a method to increase the efficiency of biogas plants that are based on anaerobic digestion of swine manure. Also, the pretreatment of wheat straw with the wet explosion method was studied and the efficiency of the wet explosion process was evaluated based on (a) the sugars release and (b) the methane potential of the pretreated wheat straw compared to that of the raw biomass. It was found that, although a high release of soluble sugars was observed after wet explosion, the methane obtained from the wet-exploded wheat straw was slightly lower compared to that from the raw biomas s. On the other hand, the results from the codigestion of raw (non-pretreated) wheat straw with swine manure were very promising, suggesting that 4.6 kg of straw added to 1t of manure increase the methane production by 10%. Thus, wheat straw can be considered as a promising, low-cost biomass for increasing the methane productivity of biogas plants that are based mainly on swine manure.

Energy balance and cost-benefit analysis of biogas production from perennial energy crops pretreated by wet oxidation.

Perennial crops need far less energy to plant, require less fertilizer and pesticides, and show a lower negative environmental impact compared with annual crops like for example corn. This makes the cultivation of perennial crops as energy crops more sustainable than the use of annual crops. The conversion into biogas in anaerobic digestion plants shows however much lower specific methane yields for the raw perennial crops like miscanthus and willow due to their lignocellulosic structure. Without pretreatment the net energy gain is therefore lower for the perennials than for corn. When applying wet oxidation to the perennial crops, however, the specific methane yield increases significantly and the ratio of energy output to input and of costs to benefit for the whole chain of biomass supply and conversion into biogas becomes higher than for corn. This will make the use of perennial crops as energy crops competitive to the use of corn and this combination will make the production of biogas from energy crops more sustainable.

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.

Effect of tryptone and ammonia on the biogas process in continuously stirred tank reactors treating cattle manure.

Two thermophilic continuously stirred tank reactors, R1 and R2, were subject to pulses of tryptone and ammonia. R1 was operated at an ammonia-N concentration of 3.0 g l(-1) and R2 was operated at an ammonia-N concentration of 1.7 g l(-1). Shock loads of tryptone (10 g l(-1), 10 g l(-1), 15 g l(-1)) had an immediate stimulating effect on methanogenesis for both reactors illustrated by significant peaks in methane production but also led to an organic overloading illustrated by a steep increase in volatile fatty acids (VFA) concentration. Three days after the pulses a second peak in acetate concentration and a decrease in methane production indicated an ammonia-inhibition of the acetoclastic methanogens. During the pulses of tryptone the performance of R1 was slightly more affected than R2. Pulses of ammonia (0.79 g l(-1) as N) resulted in a decrease in methane production of both reactors but no immediate increases in VFA concentrations was observed illustrating that the ammonia inhibition during this experiment was an overall inhibition of the biogas process and not only an inhibition of the methanogens.

Identifiability study of the proteins degradation model, based on ADM1, using simultaneous batch experiments.

The objective of the present study is to analyse kinetic and stoichiometric parameter values of gelatine anaerobic degradation at thermophilic range, based on an experiment designed to elucidate if volatile fatty acids (VFA) are inhibitors of the hydrolysis process. Results showed that VFA are not inhibiting the hydrolysis process. The ADM1 model adequately expressed the consecutive steps of hydrolysis and acidogenesis, with estimated kinetic values corresponding to a fast acidogenesis and slower hydrolysis. The hydrolysis was found to be the rate limiting step of anaerobic degradation. Estimation of yield coefficients based on the relative initial slopes of VFA profiles obtained in a simple batch experiment produced satisfactory results. From the identification study, it was concluded that it is possible to determine univocally the related kinetic parameter values for protein degradation if the evolution of amino acids is measured in simultaneous batch experiments, with different initial protein and amino acids concentrations.

Status of ADSW 2005.

Abstract The status of the recent developments in anaerobic digestion of solid waste (ADSW) is outlined on the basis of a selection of papers presented at the 4th International Symposium on ADSW 2005.

Strategies for the anaerobic digestion of the organic fraction of municipal solid waste: an overview.

Different process strategies for anaerobic digestion of the organic fraction of municipal solid waste (OFMSW) are reviewed weighing high-solids versus low-solids, mesophilic versus thermophilic and single-stage versus multi-stage processes. The influence of different waste characteristics such as composition of biodegradable fractions, C:N ratio and particle size is described. Generally, source sorting of OFMSW and a high content of food waste leads to higher biogas yields than the use of mechanically sorted OFMSW. Thermophilic processes are more efficient than mesophilic processes in terms of higher biogas yields at different organic loading rates (OLR). Highest biogas yields are achieved by means of wet thermophilic processes at OLRs lower than 6 kg-VS x m(-3) d(-1). High-solids processes appear to be relatively more efficient when OLRs higher than 6 kg-VS x m(-3)d(-1) are applied. Multi-stage systems show in some investigations a higher reduction of recalcitrant organic matter compared to single-stage systems, but they are seldom applied in full-scale. An extended cost-benefit calculation shows that the highest overall benefit of the process is achieved at an OLR that is lower and a hydraulic retention time (HRT) that is longer than those values of OLR and HRT, at which the highest biogas production is achieved.

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.

Thermophilic anaerobic fermentation of olive pulp for hydrogen and methane production: modelling of the anaerobic digestion process.

The present study investigates the thermophilic biohydrogen and methane production from olive pulp, which is the semi-solid residue coming from the two-phase processing of olives. It focussed on: a) production of methane from the raw olive pulp; b) anaerobic bio-production of hydrogen from the olive pulp; c) subsequent anaerobic treatment of the hydrogen-effluent with the simultaneous production of methane; and d) development of a mathematical model able to describe the anaerobic digestion of the olive pulp and the effluent of hydrogen producing process. Both continuous and batch experiments were performed. The hydrogen potential of the olive pulp amounted to 1.6 mmole H2 per g TS. The methane potential of the raw olive pulp and hydrogen-effluent was as high as 19 mmole CH4 per g TS suggesting that: a) olive pulp is a suitable substrate for methane production; and b) biohydrogen production can be very efficiently coupled with a subsequent step for methane production.

Thermal pre-treatment of primary and secondary sludge at 70 degrees C prior to anaerobic digestion.

In general, mesophilic anaerobic digestion of sewage sludge is more widely used compared to thermophilic digestion, mainly because of the lower energy requirements and higher stability of the process. However, the thermophilic anaerobic digestion process is usually characterised by accelerated biochemical reactions and higher growth rate of microorganisms resulting in an increased methanogenic potential at lower hydraulic retention times. Furthermore, thermal pre-treatment is suitable for the improvement of stabilization and could be realized at relatively low cost especially at low temperatures. The present study investigates the effect of the pre-treatment at 70 degrees C on thermophilic (55 degrees C) anaerobic digestion of primary and secondary sludge in continuously operated digesters. Thermal pre-treatment of primary and secondary sludge at 70 degrees C enhanced the removal of organic matter and the methane production during the subsequent anaerobic digestion step at 55 degrees C. It also greatly contributed to the destruction of pathogens present in primary sludge. Finally it results in enhanced microbial activities of the subsequent anaerobic step suggesting that the same efficiencies in organic matter removal and methane recovery could be obtained at lower HRTs.

Potential for biohydrogen and methane production from olive pulp.

The present study investigates the potential for thermophilic biohydrogen and methane production from olive pulp, which is the semi-solid residue coming from the two-phase processing of olives. It focussed on: a) production of methane from the raw olive pulp, b) anaerobic bio-production of hydrogen from the olive pulp, and c) subsequent anaerobic treatment of the hydrogen-effluent with the simultaneous production of methane. Both continuous and batch experiments were performed. The hydrogen potential of the olive pulp amounted to 1.6 mmole H2 per g TS. The methane potential of the raw olive pulp and hydrogen-effluent was as high as 19 mmole CH4 per g TS. This suggests that olive pulp is an ideal substrate for methane production and it shows that biohydrogen production can be very efficiently coupled with a subsequent step for methane production.

Wet oxidation treatment of organic household waste enriched with wheat straw for simultaneous saccharification and fermentation into ethanol.

Organic municipal solid waste enriched with wheat straw was subjected to wet-oxidation as a pre-treatment for subsequent enzymatic conversion and fermentation into bio-ethanol. The effect of temperature (185-195 degrees C), oxygen pressure (3-12 bar) and sodium carbonate (0-2 g l(-1) ) addition on enzymatic cellulose and hemicellulose convertibility was studied at a constant wet oxidation retention time of 10 minutes. An enzyme convertibility assay at high enzyme loading (25 filter paper unit (FPU) g(-1) dry solids (DS) added) showed that up to 78% of the cellulose and up to 68% of the hemicellulose in the treated waste could be converted into respectively hexose and pentose sugars compared to 46% for cellulose and 36% for hemicellulose in the raw waste. For all wet oxidation conditions tested, total carbohydrate recoveries were high (> 89%) and 44-66% of the original lignin could be converted into non-toxic carboxylic acids mainly (2.2-4.5 % on DS basis). Simultaneous saccharification and fermentation (SSF) of the treated waste at 10% DS by Saccharomyces cerevisae yielded average ethanol concentrations of 16.5 to 22 g 1(-1) for enzyme loadings of 5 and 25 FPU g(-1) DS, respectively. The cellulose to ethanol conversion efficiency during SSF was 50, 62, 65 and 70% for a total enzyme loading of 5, 10, 15 and 25 FPU g(-1) DS, respectively. Hence, this study shows that wet oxidation is a suitable pre-treatment for the conversion of organic waste carbohydrates into ethanol and that compatible conversion yields (60-65%) can be achieved at moderate enzyme loadings.

Inhibition of ethanol-producing yeast and bacteria by degradation products produced during pre-treatment of biomass.

An overview of the different inhibitors formed by pre-treatment of lignocellulosic materials and their inhibition of ethanol production in yeast and bacteria is given. Different high temperature physical pre-treatment methods are available to render the carbohydrates in lignocellulose accessible for ethanol fermentation. The resulting hydrolyzsates contain substances inhibitory to fermentation-depending on both the raw material (biomass) and the pre-treatment applied. An overview of the inhibitory effect on ethanol production by yeast and bacteria is presented. Apart from furans formed by sugar degradation, phenol monomers from lignin degradation are important co-factors in hydrolysate inhibition, and inhibitory effects of these aromatic compounds on different ethanol producing microorganisms is reviewed. The furans and phenols generally inhibited growth and ethanol production rate (Q(EtOH)) but not the ethanol yields (Y(EtOH)) in Saccharomyces cerevisiae. Within the same phenol functional group (aldehyde, ketone, and acid) the inhibition of volumetric ethanol productivity was found to depend on the amount of methoxyl substituents and hence hydrophobicity (log P). Many pentose-utilizing strains Escherichia coli, Pichia stipititis, and Zymomonas mobilis produce ethanol in concentrated hemicellulose liquors but detoxification by overliming is needed. Thermoanaerobacter mathranii A3M3 can grow on pentoses and produce ethanol in hydrolysate without any need for detoxification.

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.

Potential for using thermophilic anaerobic bacteria for bioethanol production from hemicellulose.

A limited number of bacteria, yeast and fungi can convert hemicellulose or its monomers (xylose, arabinose, mannose and galactose) into ethanol with a satisfactory yield and productivity. In the present study we tested a number of thermophilic enrichment cultures, and new isolates of thermophilic anaerobic bacterial strains growing optimally at 70-80 degrees C for their ethanol production from D-xylose. The new isolates came from different natural and man-made systems such as hot springs, paper pulp mills and brewery waste water. The test was composed of three different steps; (i) test for conversion of D-xylose into ethanol; (ii) test for viability and ethanol production in pretreated wheat straw hemicellulose hydrolysate; (iii) test for tolerance against high D-xylose concentrations. A total of 86 enrichment cultures and 58 pure cultures were tested and five candidates were selected which successfully fulfilled the criteria defined for the screening test.