Methanol based elongation of propionate and propionate and acetate in continuous and batch mixed culture systems
Smit, Sanne de; Leeuw, K.D. de; Buisman, C.J.N. ; Strik, D.P.B.T.B. - \ 2019
chain elongation - selective pressure - open-culture fermentation - mixed culture fermentation - biobased chemicals - methanol - butyrate - n-valerate
Continuous methanol based chain elongation of propionate to n-valerate was performed in a mixed culture reactor. Additionally, the study shows simultaneous methanol based elongation of propionate and acetate to respectively n-valerate and iso/n-butyrate in a continuous mixed culture reactor. A range of mixed culture batch experiments was performed with methanol and propionate and the initial pH ranging from 5 to 7.5 (steps of 0.5)
Microbial chain elongation based on methanol
Chen, Wei-Shan - \ 2017
Wageningen University. Promotor(en): C.J.N. Buisman; C. Kroeze, co-promotor(en): D.P.B.T.B. Strik. - Wageningen : Wageningen University - ISBN 9789463431989 - 201
feedstocks - renewable resources - organic wastes - waste utilization - fermentation - methanol - industriële grondstoffen - vervangbare hulpbronnen - organisch afval - afvalhergebruik - fermentatie - methanol
Our society relies heavily on fossil resources to fulfill our energy and commodity demands and this dependence has led to negative economic, environmental and societal consequences. The re-generation rate of fossil resources is much slower than their consumption rate, making these resources a non-renewable feedstock for the supply of energy and goods to our society. Moreover, the rapid consumption of fossil resources releases the carbon sequestrated in the last few million years in a much shorter time span, which contributes to the carbon dioxide (CO2) concentration increase in the atmosphere and potentially global warming. The geographically-uneven distribution of fossil resources also induces social insecurities and political conflicts. An alternative feedstock is necessary for energy and goods supply to our society, and such alternative feedstock should be renewable, economically sustainable, environmentally sound and geographically wide-spread,.
Organic waste is an emerging and promising alternative feedstock. The production of organic waste is inevitable, occurs in large quantities and is geographically wide-spread, especially the so-called “mixed organic waste,” e.g. organic fraction of municipal solid waste (OFMSW) and food processing waste. Mixed organic waste contains a large quantity of carbon materials that can be valorised into energy carriers and commodities. However, the extremely heterogeneous composition and the relatively high water content of mixed organic waste make its valorisation via the current waste management methods (e.g. incineration, composting and anaerobic digestion) less efficient and not economically attractive. Given this context, a novel bioprocess based on a mixed culture fermentation, i.e. microbial chain elongation, was developed to promote the valorisation of mixed organic waste. In microbial chain elongation, the diverse, complex organic matter in mixed organic waste are homogenised via hydrolysis and bacterial acidification into basic building blocks; like short chain fatty acids (SCFAs), CO2 and hydrogen (H2). After the homogenisation, energy-rich co-substrates like ethanol are added to these basic building blocks to synthesise medium chain fatty acids (MCFAs) via a mixed culture fermentation. MCFAs are organic compounds with a higher economic value and a higher energy content. Microbial chain elongation can be operated under a non-sterile condition, which makes it applicable to valorise mixed organic waste where diverse microorganisms exist. Caproate is the most dominant product in the microbial chain elongation of mixed organic waste and ethanol, which can be produced at a high rate and selectivity. Caproate has a higher economic value, a lower solubility in water and an interesting market potential. Thus, caproic acid production from mixed organic waste and ethanol via microbial chain elongation is currently undergoing up-scaling and commercialisation.
Many studies were done to improve the process of caproate production via microbial chain elongation to make it of industrial interest. The on-going commercialisation of microbial chain elongation also supports the economic feasibility. However, until now, no study addressed the environmental sustainability of microbial chain elongation. Chapter 2 of this thesis took the first attempt in analysing the life-cycle environmental impacts of caproic acid production from organic waste via microbial chain elongation, based on the literature and existing business case. The use of ethanol as a co-substrate (i.e. the electron donor) was shown to be the largest cause the environmental impact. This was found in in all assessed cases and all impact categories studied, and regardless of the feedstocks from which ethanol was produced. An alternative for ethanol as electron donor in microbial chain elongation is, therefore, an effective way to improve the environmental sustainability of microbial chain elongation.
In Chapter 3, we investigated the use of methanol as an alternative electron donor in microbial chain elongation, i.e. methanol chain elongation, for butyrate and caproate production. Methanol chain elongation was previously demonstrated using a pure culture, but never with a mixed culture. To employ organic waste as feedstock, the feasibility of applying methanol chain elongation in an open mixed culture condition needs to be investigated. In Chapter 3, it was demonstrated in a batch incubation that methanol chain elongation could occur with a mixed culture, where butyrate was the dominant product (4.2 g/L). Caproate production via methanol chain elongation was also demonstrated, though only in a low concentration (0.1 g/L). In a continuous reactor operation, continuous butyrate production (1.5 g/L.day) was achieved via microbial chain elongation of acetate and methanol. However, caproate was not observed in the continuous methanol chain elongation. Interestingly, microorganisms that can perform methanol chain elongation were likely present in the inoculum taken from a previous ethanol chain elongation reactor without any methanol supplement.
In Chapter 4, the use of methanol chain elongation to synthesise a novel product, i.e. isobutyrate, was proposed and investigated. Methanol chain elongation was found to continuously produce butyrate as the main metabolite, the accumulation of which was found to trigger isobutyrate formation in several previous methanogenic anaerobic digestion studies. It was, therefore, hypothesised that by elevating the butyrate concentration in the medium, methanol chain elongation might be able to produce isobutyrate as another metabolite. The result showed that isobutyrate could be produced as the main product, up to 6.2 g/L, when using acidified supermarket food waste and methanol as the substrate. A continuous methanol chain elongation using synthetic medium was also performed, which achieved a production rate of 2.0 g/L.day over five hydraulic retention times. Moreover, the production of isovalerate was also observed. Isobutyrate has a much larger market potential than caproate, though its production relies wholly on fossil-based feedstock. Isobutyrate biosynthesis was demonstrated in previous studies, but was only achieved using metabolically engineered microorganisms as the biocatalyst and glucose as the substrate. Methanol chain elongation, in contrast, could employ derivatives from organic waste as the substrates and a self-regenerating mixed culture biocatalyst for producing isobutyrate. Moreover, methanol chain elongation may be integrated into the current microbial chain elongation production facility without a significant infrastructure retrofit. All these advantages make methanol chain elongation an interesting and promising isobutyrate production process. The relatively large market potential of isobutyrate promotes the application of chain elongation and the use of organic waste for value-added chemical production.
In Chapter 5, isobutyrate production was integrated with the caproate production via microbial chain elongation, by concurrently feeding both methanol and ethanol to a mixed culture. The result from Chapter 3 supports the possibility of coexistence of ethanol and methanol chain elongation microorganisms in the same microbiome. In Chapter 4, the possible concurrence of methanol and ethanol chain elongation was also observed. Based on these observations, we hypothesised that methanol and ethanol chain elongation could be integrated to simultaneously produce caproate and isobutyrate. The result showed that such integration was possible when a stable pH was maintained. When pH was controlled between 6.2 – 6.5 and butyrate was supplied in the medium, caproate and isobutyrate could be produced simultaneously. Additionally, increasing the ethanol feeding rate promoted the chain elongation of butyrate to caproate via ethanol chain elongation. The outcome of this chapter demonstrated the possibility of producing two valuable products in a single reactor with a mixed culture which, coupled with further process improvement, may be of industrial interest.
In Chapter 6, we reflected on the caproate production performance of methanol chain elongation, in comparison with other electron donors used in microbial chain elongation, i.e. ethanol and lactate. Furthermore, we also reflected on the isobutyrate production via methanol chain elongation, in comparison with other emerging products in microbial chain elongation. These reflections could serve as a benchmark for methanol chain elongation as a waste management strategy. Based on this benchmarking, we proposed that methanol chain elongation is a promising bioprocess for isobutyrate production but not for caproate production. A potential strategy for improving the isobutyrate production via methanol chain elongation was proposed and discussed. The outcomes of this thesis may contribute to future application and assessments of microbial chain elongation in waste management. It may fuel discussion on how to further promote microbial chain elongation for a more sustainable waste management.
EnAlgae Decision Support Toolset: model validation
Kenny, Philip ; Visser, Chris de; Skarka, Johannes ; Sternberg, Kirstin ; Schipperus, Roelof ; Silkina, Alla ; Ginnever, Naomi - \ 2015
Swansea : Swansea University - 25
biobased economy - bioenergy - biomass - algae culture - algae - bioethanol - biodiesel - methanol - seaweeds - seaweed culture - biobased economy - bio-energie - biomassa - algenteelt - algen - bioethanol - biodiesel - methanol - zeewieren - zeewierenteelt
One of the drivers behind the EnAlgae project is recognising and addressing the need for increased availability of information about developments in applications of algae biotechnology for energy, particularly in the NW Europe area, where activity has been less intense than in other areas of the globe. Such information can be of benefit in coordinating research activities, stimulating targeted investment to develop promising technologies and to guide key policy decisions. To make this a reality, EnAlgae has developed a Decision Support Toolset (DST) to enable improved evaluation of state of the art algal biotechnology and to compare alternative routes to utilising algal biomass.
One-carbon metabolism in acetogenic and sulfate-reducing bacteria
Visser, M. - \ 2015
Wageningen University. Promotor(en): Fons Stams. - Wageningen : Wageningen University - ISBN 9789462571730 - 210
anaërobe microbiologie - metabolisme - koolmonoxide - methanol - alcohol dehydrogenase - sulfaat reducerende bacteriën - genetische analyse - eiwitexpressieanalyse - anaerobic microbiology - metabolism - carbon monoxide - methanol - alcohol dehydrogenase - sulfate reducing bacteria - genetic analysis - proteomics
One-carbon metabolism in acetogenic and sulfate-reducing bacteria
Life on earth is sustained by the constant cycling of six essential elements: oxygen, hydrogen, nitrogen, sulfur, phosphorous, and carbon. The continuous cycling of these elements is due to geo-chemical processes and the combined metabolism of all life on earth. Microorganisms like bacteria and archaea play a major role in this. This is also true for the carbon cycle. In this cycle carbon dioxide and methane are two important C-1 compounds present in the atmosphere. Carbon dioxide is the highest oxidative state of carbon while methane is the highest reduced form of carbon. The art to use light to produce organic compounds and conserve energy from the highest oxidative state of carbon is called photosynthesis and is performed by plants, algae and cyanobacteria. Photosynthesis is not the only system to fix carbon from carbon dioxide. Chemolithotrophs can fix carbon from carbon dioxide using inorganic electron donors, like hydrogen. Subsequently, fixed carbon can be used by other organisms, which also makes life possible for them. Microorganisms play a major role in the degradation of complex organic matter, producing smaller compounds including C-1 compounds. C-1 compounds other than carbon dioxide are e.g. carbon monoxide (CO), methanol and formate. Bacteria and archaea can utilize these relative simple compounds in the presence and absence of oxygen, alone and in cooperation with others (syntrophy). The complex and simple carbon compounds are finally oxidized to carbon dioxide, which closes the carbon cycle.
In addition to their importance to the carbon cycle, one carbon compounds like CO, methanol and formate are important for several applications. They are used as a building block for the production of chemicals. They are also used for bioremediation purposes and for wastewater treatment. Therefore, it is important to gain insight in the one carbon metabolism of microorganisms. The research described in this thesis focuses on the proteins and encoding genes involved in anaerobic degradation of C1 compounds by using genome and proteome analysis.
In Chapter 2 the genomes of two closely related sulfate-reducing bacteria, Desulfotomaculum nigrificans and D. nigrificans strain CO-1-SRB, are compared including their CO metabolism. Both the D. nigrificans type strain and strain CO-1-SRB can grow with CO. However, there are differences. The type strain can grow with 20% CO coupled to sulfate reduction in the presence of yeast extract, while strain CO-1-SRB can grow with 100% CO in the presence of yeast extract. Moreover, strain CO-1-SRB can grow with CO in the presence and absence of sulfate. It couples the oxidation of CO to carbon dioxide to hydrogen production. This conversion, the protein complex involved, and the genes coding for these proteins have been described before in other microorganisms. The genome of strain CO-1-SRB contains the genes coding for this protein complex while the genome of the D. nigrificans type strain does not. However, the genome of the type strain contains genes encoding two other CO dehydrogenases. This indicates that one or both are necessary for the type strain to grow with 20% CO. Additional research on the different CO dehydrogenases and their regulation is essential to assess if all different CO dehydrogenases can facilitate growth and how they are linked to for example creating a proton motive force for ATP production.
The methanol metabolism of anaerobic bacteria seems to differ more from that of methanogens than initially described. Methanogens use a methanol methyltransferase system that consists of two methyltranferases, methyltransferase 1 (subunits MtaB and MtaC) and methyltransferase 2 (MtaA). The methyl group from methanol is transferred to the MtaC subunit by MtaB. Subsequently, MtaA transports the methyl group from MtaC to coenzyme M. A genome and proteome analysis of the acetogenic bacterium Sporomusa strain An4 suggests that instead of MtaA a methyl-tetrahydrofolate methyltransferase is involved in the transport of the methyl bound to MtaC to tetrahydrofolate (Chapter 3).
Research done on the methanol metabolism of the sulfate-reducing bacterium Desulfotomaculum kuznetsovii also shows differences with that of methanogens (Chapter 5). The methanol methyltransferase system is vitamin B12 and cobalt dependent. D. kuznetsovii grows with methanol and sulfate, but can do this in presence and absence of vitamin B12 and cobalt. In the absence of vitamin B12 and cobalt D. kuznetsovii grows slower and reaches a lower optical density compared to growth in the presence of vitamin B12 and cobalt. This suggests that D. kuznetsovii can use both a methyltransferase system and a vitamin B12 and cobalt independent system for the degradation of methanol. Proteome results confirm this and suggest that the vitamin B12 and cobalt independent system consists of an alcohol dehydrogenase and an aldehyde ferredoxin oxidoreductase. Moreover, the alcohol dehydrogenase seems to be involved in the oxidation of both methanol and ethanol (Chapter 5). The presence of two methanol degradation pathways give an ecological advantage to D. kuznetsovii in environments containing methanol and sulfate but limiting cobalt and vitamin B12 concentrations. Future research should elucidate if more sulfate-reducing bacteria, or perhaps even acetogenic bacteria, have two methanol degrading pathways.
Additional to the genome analysis of D. kuznetsovii to assess the genes coding for the proteins involved in the two methanol degradation pathways, the genome was also analyzed to assess genes encoding other degradation pathways (Chapter 4). This analysis shows many genes present in D. kuznetsovii are also present in Pelotomaculum thermopropionicum. P. thermopropionicum is known to degrade propionate in syntrophic interaction with a methanogen. D. kuznetsovii can also degrade propionate, but only coupled to sulfate reduction and not in syntrophy with methanogens. Moreover, P. thermopropionicum is not able to reduce sulfate. D. kuznetsovii is the only close related, non-syntrophic, propionate degrader of which the genome is available. Therefore, a genome comparison was performed between D. kuznetsovii and P. thermopropionicum to define the differences between a non-syntrophic and a syntrophic lifestyle. D. kuznetsovii misses membrane bound protein complexes like hydrogenases and an extra-cytoplasmic formate dehydrogenase. In order to expand the analysis between non-syntrophs and syntrophs, more genomes of propionate- and butyrate-degrading bacteria were included (Chapter 6). This extended analysis shows that the genomes of non-syntrophs do not contain genes coding for an extra-cytoplasmic formate dehydrogenase, in contrast to all syntrophs included in the analysis. This indicates the importance of this protein complex and the importance of formate as an interspecies electron carrier in syntrophic degradation of propionate and butyrate. Thanks to the extra cytoplasmic formate dehydrogenase the syntrophic bacteria can couple the degradation of propionate and butyrate to formate production. Subsequently, the formate is utilized by methanogens to produce methane. This keeps the formate concentration low, which is necessary for the entire process to be energetically favorable.
Base-Free, One-Pot Chemocatalytic Conversion of Glycerol to Methyl Lactate using Supported Gold Catalysts
Purushothaman, R.K.P. ; Haveren, J. van; Melian-Cabrera, I. ; Eck, E.R.H. van; Heeres, H.J. - \ 2014
ChemSusChem 7 (2014)4. - ISSN 1864-5631 - p. 1140 - 1147.
lactic-acid - selective oxidation - commodity chemicals - aqueous-solutions - d-glucose - hydrogenolysis - transformation - methanol - alcohol - insight
We report an efficient one-pot conversion of glycerol (GLY) to methyl lactate (MLACT) in methanol in good yields (73% at 95% GLY conversion) by using Au nanoparticles on commercially available ultra-stable zeolite-Y (USY) as the catalyst (160 degrees C, air, 47bar pressure, 0.25M GLY, GLY-to-Au mol ratio of 1407, 10h). The best results were obtained with zeolite USY-600, a catalyst that has both Lewis and BrOnsted sites. This methodology provides a direct chemo-catalytic route for the synthesis of MLACT from GLY. MLACT is stable under the reaction conditions, and the Au/USY catalyst was recycled without a decrease in the activity and selectivity.
Genome analysis of Desulfotomaculum kuznetsovii strain 17T reveals a physiological similarity with Pelotomaculum thermopropionicum SIT
Visser, M. ; Worm, P. ; Muyzer, G. ; Pereira, I.A.C. ; Schaap, P.J. ; Plugge, C.M. ; Kuever, J. ; Parshina, S.N. ; Nazina, T.N. ; Ivanova, A.E. ; Bernier-Latmani, R. ; Goodwin, L.A. ; Kyrpides, N. ; Woyke, T. ; Chain, P. ; Davenport, K.W. ; Spring, S. ; Klenk, H.P. ; Stams, A.J.M. - \ 2013
Standards in Genomic Sciences 8 (2013)1. - ISSN 1944-3277 - p. 69 - 87.
sulfate-reducing bacteria - clostridium-thermoaceticum - carbon-monoxide - gen. nov. - sequence - prediction - biosynthesis - oxidation - methanol - enzymes
Desulfotomaculum kuznetsovii is a moderately thermophilic member of the polyphyletic spore-forming genus Desulfotomaculum in the family Peptococcaceae. This species is of interest because it originates from deep subsurface thermal mineral water at a depth of about 3000 m. D. kuznetsovii is a rather versatile bacterium as it can grow with a large variety of organic substrates, including short-chain and long-chain fatty acids, which are degraded completely to carbon dioxide coupled to the reduction of sulfate. It can grow methylotrophically with methanol and sulfate and autotrophically with H2 + CO2 and sulfate. For growth it does not require any vitamins. Here, we describe the features of D. kuznetsovii together with the genome sequence and annotation. The chromosome has 3,601,386 bp organized in one contig. A total of 3567 candidate protein-encoding genes and 58 RNA genes were identified. Genes of the acetyl-CoA pathway possibly involved in heterotrophic growth with acetate and methanol, and in CO2 fixation during autotrophic growth are presented. Genomic comparison revealed that D. kuznetsovii shows a large similarity with Pelotomaculum thermopropionicum. Genes involved in propionate metabolism of these two strains show a strong similarity. However, main differences are found in genes involved in the electron acceptor metabolism
Sulfate reduction during the acidification of sucrose at pH 5 under thermophilic (55 degrees C) conditions. I: Effect of trace metals
Lopes, S.I.C. ; Capela, M.I. ; Lens, P.N.L. - \ 2010
Bioresource Technology 101 (2010)12. - ISSN 0960-8524 - p. 4269 - 4277.
anaerobic granular sludge - desulfovibrio-desulfuricans - waste-water - inhibition - speciation - toxicity - degradation - 8-degrees-c - reactors - methanol
This work studied the effect of supplying trace metals (7.5 mu M Fe and 0.5 mu M Co, Ni, Mn, Zn, Cu, B, Se, Mo and W) on sulfate reduction and acidification in thermophilic (55 degrees C) UASB reactors fed with sucrose (4 gCOD (I-reactor d)(-1)) operated at a reactor mixed liquor pH controlled at 5. Trace metals were supplied to one UASB reactor and were omitted from the influent of a second UASB reactor. The influence of different trace metal concentrations was further assessed in batch tests performed with the sludge from the UASB reactor receiving no trace metals. The absence of trace metals in the influent did not affect the performance of the acidifying UASB reactor throughout the 305 day long reactor run, but supplying low concentrations of trace metals inhibited sulfate reduction.
Synthesis of biobased N-methylpyrrolidone by one-pot cyclization and methylation of c-aminobutyric acid
Lammens, T.M. ; Franssen, M.C.R. ; Scott, E.L. ; Sanders, J.P.M. - \ 2010
Green Chemistry 12 (2010)8. - ISSN 1463-9262 - p. 1430 - 1436.
dimethyl carbonate - faujasites - chemistry - methanol
N-Methylpyrrolidone (NMP) is an industrial solvent that is currently based on fossil resources. In order to prepare it in a biobased way, the possibility to synthesize NMP from -aminobutyric acid (GABA) was investigated, since GABA can be obtained from glutamic acid, an amino acid that is present in many plant proteins. Cyclization of GABA to 2-pyrrolidone and subsequent methylation of 2-pyrrolidone to NMP was achieved in a one-pot procedure, using methanol as the methylating agent and a halogen salt (i.e. ammonium bromide) as a catalyst. A selectivity above 90% was achieved, as well as a high conversion. Methylation of 2-pyrrolidone could also be done with dimethyl carbonate, but then the selectivity for NMP was less (67%).
Endogenous and bioaugmented sulphate reduction in calcareous gypsiferous soils
Alfaya, F. ; Cuenca-Sanchez, M. ; Garcia-Orenes, F. ; Lens, P.N.L. - \ 2009
Environmental Technology 30 (2009)12. - ISSN 0959-3330 - p. 1305 - 1312.
sludge bed reactors - granular sludge - reducing bacteria - anaerobic reactor - electron-donors - methanol - bioreactor - acetate - ethanol - sucrose
Gypsiferous soils have a high agricultural value, but their utilization is limited by the presence of gypsum that can induce hardpan and vertical crusting. This paper reports on sulphate reduction in this soil type as a basis of a bioremediation technology to remove the gypsum content of calcareous gypsiferous soils. Both the endogenous and bioaugmented sulphate-reducing potential of the soil was assessed in anaerobic batch tests. An endogenous population of sulphate-reducing bacteria was found to be present in the calcareous gypsiferous soil investigated, which could form the basis of both in situ and ex situ bioremediation schemes for this soil type.
Atypical one-carbon metabolism of an acetogenic and hydrogenogenic Moorella thermoacetica strain
Jiang, B. ; Henstra, A.M. ; Paulo, P.L. ; Balk, M. ; Doesburg, W.C.J. van; Stams, A.J.M. - \ 2009
Archives of Microbiology 191 (2009)2. - ISSN 0302-8933 - p. 123 - 131.
performance liquid-chromatography - clostridium-thermoaceticum - sp-nov. - carboxydothermus-hydrogenoformans - thermophilic bacterium - gen. nov. - methanol - thermoautotrophicum - dna - purification
A thermophilic spore-forming bacterium (strain AMP) was isolated from a thermophilic methanogenic bioreactor that was fed with cobalt-deprived synthetic medium containing methanol as substrate. 16S rRNA gene analysis revealed that strain AMP was closely related to the acetogenic bacterium Moorella thermoacetica DSM 521T (98.3% sequence similarity). DNA¿DNA hybridization showed 75.2 ± 4.7% similarity to M. thermoacetica DSM 521T, suggesting that strain AMP is a M. thermoacetica strain. Strain AMP has a unique one-carbon metabolism compared to other Moorella species. In media without cobalt growth of strain AMP on methanol was only sustained in coculture with a hydrogen-consuming methanogen, while in media with cobalt it grew acetogenically in the absence of the methanogen. Addition of thiosulfate led to sulfide formation and less acetate formation. Growth of strain AMP with CO resulted in the formation of hydrogen as the main product, while other CO-utilizing Moorella strains produce acetate as product. Formate supported growth only in the presence of thiosulfate or in coculture with the methanogen. Strain AMP did not grow with H2/CO2, unlike M. thermoacetica (DSM 521T). The lack of growth with H2/CO2 likely is due to the absence of cytochrome b in strain AMP
Cobalt toxicity in anaerobic granular sludge: influence of chemical speciation
Bartacek, J. ; Fermoso, F.G. ; Baldo-Urrutia, A.M. ; Hullebusch, E.D. van; Lens, P.N.L. - \ 2008
Journal of Industrial Microbiology and Biotechnology 35 (2008)11. - ISSN 1367-5435 - p. 1465 - 1474.
donnan membrane technique - metal-ion concentrations - trace-metals - sequential extraction - organic-matter - biotic ligand - heavy-metals - methanol - bioavailability - deprivation
The influence of cobalt speciation on the toxicity of cobalt to methylotrophic methanogenesis in anaerobic granular sludge was investigated. The cobalt speciation was studied with three different media that contained varying concentrations of complexing ligands [carbonates, phosphates and ethylenediaminetetraacetic acid (EDTA)]. Three fractions (nominal added, dissolved and free) of cobalt were determined in the liquid media and were correlated with data from batch toxicity experiments. The average concentration of cobalt that was required for 50% inhibition of methanogenic activity (IC50) for free Co2+ in the three sets of measurements was 13 mu mol/L with a standard deviation of 22% and a similarity of 72% between the data obtained in the three different media for the range of cobalt concentrations investigated. The standard deviation of the IC50 for the other two fractions was much higher, i.e. 85 and 144% for the added cobalt and dissolved cobalt, respectively, and the similarity was almost 0% for both fractions. Complexation (and precipitation) with EDTA, phosphates and carbonates was shown to decrease the toxicity of cobalt on methylotrophic methanogenesis. The free cobalt concentration is proposed to be the key parameter to correlate with cobalt toxicity. Thus, the toxicity of cobalt to granular sludge can be estimated based on the equilibrium-free cobalt concentration.
Effect of hydraulic retention time on sulfate reduction in a carbon monoxide fed thermophilic gas lift reactor
Sipma, J. ; Osuna, M.B. ; Lettinga, G. ; Stams, A.J.M. ; Lens, P.N.L. - \ 2007
Water Research 41 (2007)9. - ISSN 0043-1354 - p. 1995 - 2003.
biological sulfate - reducing bacteria - conversion - growth - energy - co - methanol - h-2
Thermophilic hydrogenogenic carbon monoxide (CO) converting microorganisms present in anaerobic sludge play a crucial role in the application of CO as electron donor for sulfate reduction. Hydrogenogenic CO conversion was investigated in a gas lift reactor (55 °C) at different hydraulic retention times (HRT). Operation at a HRT>9 h resulted in predominant consumption of CO-derived H2 by methanogens (up to 90%) and thus in a poor sulfate reduction efficiency of less than 15%. At HRTs5.5 h resulted in a dominance of methanogenesis over sulfate reduction. The sulfate reduction rates were limited by the amount of CO supplied and its conversion efficiency (about 85%) at higher CO loads likely resulting from a low biomass retention.
Polylactide films formed by immersion precipitation: Effects of additives, nonsolvent, and temperature
Sawalha, H.I.M. ; Schroën, C.G.P.H. ; Boom, R.M. - \ 2007
Journal of Applied Polymer Science 104 (2007)2. - ISSN 0021-8995 - p. 959 - 971.
induced phase-separation - pvdf membrane formation - asymmetric membranes - systems - water - morphology - methanol - transitions - mechanism - behavior
The influence of nonsolvent, crystallinity of the polymer film, and addition of dodecane (a poor solvent for the polymer and for the nonsolvent) on the morphology of polylactides films has been investigated and was related to phase separation behavior. Both amorphous poly-DL-lactide (PDLLA) and crystalline poly-L-lactide (PLLA) were dissolved in dichloromethane, and subsequently films were made by immersion in nonsolvent baths. PDLLA gave dense films without any internal structure, since the structure was not solidified by crystallization or glassification. PLLA films show varying structure depending on the nonsolvent. With methanol, asymmetric morphologies were observed as a result from combined liquid-liquid demixing and crystallization, while with water symmetric spherulitic structures were formed. As a next step, dodecane was added, which is not miscible with the nonsolvent, and we found it to have a strong influence on the morphology of the films. The PDLLA films with dodecane did not collapse: a closed cell structure was obtained. In PLLA films, dodecane speeds up phase separation and induces faster crystallization in the films, and the porosity, size of the pores, and interconnectivity increased. When the PLLA solutions were subjected to a heat pretreatment, crystallization could be postponed, which yielded a cellular structure around dodecane, which did not contain spherulites anymore
Effect of Temperature and High Pressure on the Activity and Mode of Action of Fungal Pectin Methyl Esterase
Duvetter, T. ; Fraeye, I. ; Sila, D.N. ; Verlent, I. ; Smout, C. ; Clynen, E. ; Schoofs, L. ; Schols, H.A. ; Hendrickx, M. ; Loey, A. van - \ 2006
Biotechnology Progress 22 (2006)5. - ISSN 8756-7938 - p. 1313 - 1320.
aspergillus-niger - apple pectin - pectinmethylesterase - firmness - endopolygalacturonase - methylesterase - infusion - methanol - enzyme - fruits
Pectin was de-esterified with purified recombinant Aspergillus aculeatus pectin methyl esterase (PME) during isothermal-isobaric treatments. By measuring the release of methanol as a function of treatment time, the rate of enzymatic pectin conversion was determined. Elevated temperature and pressure were found to stimulate PME activity. The highest rate of PME-catalyzed pectin de-esterification was obtained when combining pressures in the range 200-300 MPa with temperatures in the range 50-55 C. The mode of pectin de-esterification was investigated by characterizing the pectin reaction products by enzymatic fingerprinting. No significant effect of increasing pressure (300 MPa) and/or temperature (50 C) on the mode of pectin conversion was detected.
Hydrogenogenic CO Conversion in a Moderately Thermophilic (55 C) Sulfate-Fed Gas Lift Reactor: Competition for CO-Derived H2
Sipma, J. ; Lettinga, G. ; Stams, A.J.M. ; Lens, P.N.L. - \ 2006
Biotechnology Progress 22 (2006)5. - ISSN 8756-7938 - p. 1327 - 1334.
carbon-monoxide conversion - anaerobic bioreactor sludges - growth - reduction - methanogenesis - methanol - energy - 2-bromoethanesulfonate - chloroform - bacteria
Thermophilic (55 °C) sulfate reduction in a gas lift reactor fed with CO gas as the sole electron donor was investigated. The reactor was inoculated with mesophilic granular sludge with a high activity of CO conversion to hydrogen and carbon dioxide at 55 °C. Strong competition for H2 was observed between methanogens and sulfate reducers, while the homoacetogens present consumed only small amounts of H2. The methanogens appeared to be more sensitive to pH and temperature shocks imposed to the reactor, but could not be completely eliminated. The fast growth rates of the methanogens (generation time of 4.5 h) enabled them to recover fast from shocks, and they rapidly consumed more than 90% of the CO-derived H2. Nevertheless, steep increases in sulfide production in periods with low methane production suggests that once methanogenesis is eliminated, sulfate reduction with CO-rich gas as electron donor has great potential for thermophilic biodesulfurization
Volatile organic sulfur compounds in anaerobic sludge and sediments: biodegradation and toxicity
Leerdam, R.C. van; Bok, F.A.M. de; Lomans, B.P. ; Stams, A.J.M. ; Lens, P.N.L. ; Janssen, A.J.H. - \ 2006
Environmental Toxicology and Chemistry 25 (2006)12. - ISSN 0730-7268 - p. 3101 - 3109.
microbiële afbraak - slib - sediment - anaërobe behandeling - afvalwaterbehandeling - sulfaten - thiolen - reductie - sulfaat - methanol - biodegradatie - microbial degradation - sludges - sediment - anaerobic treatment - waste water treatment - sulfates - thiols - reduction - sulfate - methanol - biodegradation - fresh-water sediments - dimethyl sulfide - membrane bioreactor - waste air - methanethiol - degradation - inhibition - removal - ph - methanogens
A variety of environmental samples was screened for anaerobic degradation of methanethiol, ethanethiol, propanethiol, dimethylsulfide, and dimethyldisulfide. All sludge and sediment samples degraded methanethiol, dimethylsulfide, and dimethyldisulfide anaerobically. In contrast, ethanethiol and propanethiol were not degraded by the samples investigated under any of the conditions tested. Methanethiol, dimethylsulfide, and dimethyldisulfide were mainly degraded by methanogenic archaea. In the presence of sulfate and the methanogenic inhibitor bromoethane sulfonate, degradation of these compounds coupled to sulfate reduction occurred as well, but at much lower rates. Besides their biodegradability, also the toxicity of methanethiol, ethanethiol, and propanethiol to methanogenesis with methanol, acetate, and H2/CO2 as the substrates was assessed. The 50% inhibition concentration of methanethiol on the methane production from these substrates ranged between 7 and 10 mM. The 50% inhibition concentration values of ethanethiol and propanethiol for the degradation of methanol and acetate were between 6 and 8 mM, whereas hydrogen consumers were less affected by ethanethiol and propanethiol, as indicated by their higher 50% inhibition concentration (14 mM). Sulfide inhibited methanethiol degradation already at relatively low concentrations: methanethiol degradation was almost completely inhibited at an initial sulfide concentration of 8 mM. These results define the operational limits of anaerobic technologies for the treatment of volatile organic sulfur compounds in sulfide-containing wastewater streams
The effect of trace elements on the metabolism of methanogenic consortia
Jiang, B. - \ 2006
Wageningen University. Promotor(en): Fons Stams. - [S.l.] : S.n. - ISBN 9789085043607 - 122
sporenelementen - metabolisme - methanol - anaërobe afbraak - methanobacteriaceae - trace elements - metabolism - methanol - anaerobic digestion - methanobacteriaceae
Trace metals are essential for the growth and metabolism of anaerobic microorganisms, duo to their roles in key enzymes or cofactors of metabolic pathways. The requirement of trace metals has been recognized. But, proper dosing of these metals in anaerobic treatment system as nutrient still is a great challenge, since dosing of a metal at a high concentration is toxic for growth of microorganisms, and dosing of a specific metal may lead out-compete of one group of microorganisms by the other. In order to obtain knowledge for optimization of metal dosing of anaerobic treatment system, the influence of trace metals, like cobalt, nickel, tungsten and molybdenum on the conversion of methanol and propionate were studied in this research.By using cobalt-sufficient medium, a methanogenic enrichment culture was enriched from a thermophilic lab-scale UASB reactor fed with methanol as carbon and energy source. From which a novel thermophilic obligate methylotrophic methanogenicarchaeon, strain L2FAW T , was isolated and characterized as Methanomethylovoransthermophila . The growth of strain L2FAW T on methanol is stimulated by the addition of cobalt; the optimal cobalt concentration is 0.5 to 2 M. therefore, cobalt is important for direct methanol conversion by this methanogen.On the other hand, a syntrophiccocultureof methanol degradation was enriched from the same sludge by using cobalt deficient medium, which consisted of a homoacetogen and ahydrogenotrophicmethanogen. Thiscoculturedegrades methanol partially to acetate and partially to methane, depending on the presence of cobalt. Acetate is the main product when cobalt is presence at high concentration; otherwise methane is formed as dominant products. Therefore, cobalt plays a role in the regulation of the pathway of methanol conversion. The optimal cobalt concentration of thecoculturefor complete methanogenesis from methanol is about 0.1 M. A thermophilic spore-forming bacteria, strain AMP, was isolated from thecoculture, and it is most closely related to Moorellathermoaceticabased on 16S rRNA analysis. Despite its high DNA-DNA homology with M.thermoacetica , strain AMP differs from M.thermoacetica on its inability to use glucose, formate and H 2 /CO 2 , and its uniquehydrogenogenicgrowth on CO. Moreover, strain AMP can grow on formate in acoculturewith ahydrogenotrophicmethanogen. It is described for the first time that a bacterium can grow on the conversion of formate to H 2 and bicarbonate provided that hydrogen is consumed by a methanogen.The effect of cobalt and nickel on the corrinoid and F430 content and on growth of Methanosarcina barkeri on methanol was studied. Cobalt and nickel limitation was achieved and competition between cobalt and nickel uptake was observed. Uptake efficiency of cobalt was high at low cobalt concentration and decreased when the cobalt concentration in the medium was increased. Corrinoid and F430 content correlated positively with the cell content of the corresponding metal, but incorporation in the corrinoid and F430 was significant less at low cell metal contents, ranging from 35% to 80% for corrinoid and 5% to 15% for F430.The trace elements tungsten and molybdenum play an essential role in the growth of anaerobic microorganisms. Depletion of tungsten and/or molybdenum in the media did not affect axenic growth of Syntrophobacterfumaroxidans onpropionate+fumarate, indicating under these conditions this organism does not have a high tungsten or molybdenum requirement. However, growth of Methanospirillumhungatei on either formate or hydrogen and carbon dioxide required tungsten, and molybdenum can replace tungsten to some extent. Growth of the Syntrophobacter-Methanospirillumcocultureon propionate is significantly affected by the addition of these two metals. Measurement of enzyme levels in cell extracts of syntrophically grown cells indicated that the levels of hydrogenaseandformatedehydrogenaseactivity were correlated with the methane formation rates by thecocultures, which suggests both hydrogen and formate play important role in syntrophic propionate oxidation.
Cultivation of high rate sulfate reducing sludge by pH-based electron donor dosage
Paulo, P.L. ; Kleerbezem, R. ; Lettinga, G. ; Lens, P.N.L. - \ 2005
Journal of Biotechnology 118 (2005)1. - ISSN 0168-1656 - p. 107 - 116.
anaerobic-digestion - high salinity - methanol - hydrogen - reactors - methanogenesis - competition - conversion - reduction
A novel self-regulating bioreactor concept for sulfate reduction is proposed aiming for high biomass concentrations and treatment capacities. The system consists of a cell suspension of sulfate reducing bacteria in a continuous stirred tank reactor (30 degrees C) fed with a mixture of both electron donor and electron acceptor (formic acid and sulfuric acid, respectively), nutrients and phosphate buffer via a pH controller. The pH rise due to sulfate reduction is balanced with dosage of the sulfate reducing substrates as acids. The reactor concept was shown to be capable of full sulfate reduction without competition for the electron donor by methanogens and acetogens. Activity assays revealed that hardly any methanogenic activity on formate was left in the suspension by the end of the continuous run (130 days). In addition, the sulfidogenic activity with formate and H2/CO2 had increased, respectively, 3.9 and 11.6 times at the end of the experimental run. The evolution of the particle size distribution of the cell suspension over time indicated that newly grown cells have the tendency to attach together in flocs or to the existing agglomerates.
Fast determination of the degree of methyl esterification of pectins by head-space GC.
Huisman, M.M.H. ; Oosterveld, A. ; Schols, H.A. - \ 2004
Food Hydrocolloids 18 (2004)4. - ISSN 0268-005X - p. 665 - 668.
ester content - methanol
A new, fast method for the quantitative analysis of methoxyl groups in pectin using head-space gas chromatography (HS-GC) has been developed. With this method, results were obtained which were in reasonable agreement with the conventional HPLC method, and the reproducibility of the measurements is high. The advantages of the HS-GC method are that only a small amount of sample (2 mg) per analysis is needed, the chromatogram shows a nice symmetrically shaped methanol peak which is very easy to integrate, the sample preparation for HS-GC is short and easy, and for soluble pectins the sample in the head space vial can also directly be used for analysis of the galacturonic acid content and the degree of acetylation.
Thermophilic (55 - 65°C) and extreme thermophilic (70 - 80°C) sulfate reduction in methanol and formate-fed UASB reactors
Vallero, M.V.G. ; Camarero, E. ; Lettinga, G. ; Lens, P.N.L. - \ 2004
Biotechnology Progress 20 (2004)5. - ISSN 8756-7938 - p. 1382 - 1392.
anaërobe behandeling - afvalwaterbehandeling - methanol - reductie - sulfaten - temperatuur - anaerobic treatment - waste water treatment - methanol - reduction - sulfates - temperature - rate anaerobic reactor - volatile fatty-acids - 55-degrees-c - sludge - degradation - conversion - bacterium - methanogenesis - competition - performance
The feasibility of thermophilic (55-65 degreesC) and extreme thermophilic (70-80 degreesC) sulfate-reducing processes was investigated in three lab-scale upflow anaerobic sludge bed (UASB) reactors fed with either methanol or formate as the sole substrates and inoculated with mesophilic granular sludge previously not exposed to high temperatures
The feasibility of thermophilic (55-65 degreesC) and extreme thermophilic (70-80 degreesC) sulfate-reducing processes was investigated in three lab-scale upflow anaerobic sludge bed (UASB) reactors fed with either methanol or formate as the sole substrates and inoculated with mesophilic granular sludge previously not exposed to high temperatures. Full methanol and formate degradation at temperatures up to, respectively, 70 and 75 degreesC, were achieved when operating UASB reactors fed with sulfate rich (COD/SO42- = 0.5) synthetic wastewater. Methane-producing archaea (MPA) outcompeted sulfate-reducing bacteria (SRB) in the formate-fed UASB reactor at all temperatures tested (65-75 degreesC). In contrast, SRB outcompeted MPA in methanol-fed UASB reactors at temperatures equal to or exceeding 65 degreesC, whereas strong competition between SRB and MPA was observed in these reactors at 55 degreesC. A short-term (5 days) temperature increase from 55 to 65 degreesC was an effective strategy to suppress methanogenesis in methanol-fed sulfidogenic UASB reactors operated at 55 degreesC. Methanol was found to be a suitable electron donor for sulfate-reducing processes at a maximal temperature of 70 degreesC, with sulfide as the sole mineralization product of methanol degradation at that temperature.