- Email: [email protected]
Study of the co-production of butanol and hydrogen by immobilizing Clostridium acetobutylicum CICC8012
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Energy Procedia 158 Energy Procedia 00(2019) (2017)1879–1884 000–000
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10th International Conference on Applied Energy (ICAE2018), 22-25 August 2018, Hong Kong, China 10th International Conference on Applied Energy (ICAE2018), 22-25 August 2018, Hong Kong, China StudyTheof15th theInternational co-production of on butanol and hydrogen Symposium District Heating and Coolingby
immobilizing Clostridium of acetobutylicum CICC8012 Study of the co-production butanol and hydrogen by * Assessing the feasibility of using the heat demand-outdoor immobilizing Clostridium Jingyun Liu, Wencan Zhou,acetobutylicum Senqing Fan , ZeyiCICC8012 Xiao * heat temperatureJingyun function for a long-term district demand forecast SchoolLiu, of Chemical Engineering, Sichuan University, Chengdu, 610065 Xiao Wencan Zhou, Senqing Fan , Zeyi
Abstract a
a a Sichuan University, c School of Chemical Engineering, 610065 I. Andrića,b,c*, A. Pina , P. Ferrão , J. Fournierb., Chengdu, B. Lacarrière , O. Le Correc
IN+ Center for Innovation, Technology and Policy Research - Instituto Superior Técnico, Av. Rovisco Pais 1, 1049-001 Lisbon, Portugal
b Abstract Three kinds of carrier materials, activated&carbon, bagasse and brick wereDaniel, used as immobilizing carriers during fermentation Veolia Recherche Innovation, 291 Avenue Dreyfous 78520 Limay, France c acetobutylicum CICC8012. Compared with cell suspended fermentation, enhanced fermentation performance was by Clostridium Département Systèmes Énergétiques et Environnement - IMT Atlantique, 4 rue Alfred Kastler, 44300 Nantes, France achieved cell fermentation, with bagasse shorter fermentation time required. During thecarriers experiments, and Three during kinds ofimmobilizing carrier materials, activated carbon, and brick were used as immobilizing duringhydrogen fermentation butanol appear acetobutylicum to be competitive events. The best fermentation performance of butanolenhanced was obtained in the case of bagassewas as by Clostridium CICC8012. Compared with cell suspended fermentation, fermentation performance immobilizing carrier (5.804g/Lcell of butanol production, 0.22g/gfermentation of yield andtime 0.44g/L/h of productivity), while the hydrogen achieved during immobilizing fermentation, with shorter required. During the experiments, hydrogenyield and was justappear 1.41 mol/mol. The highest hydrogen (402mL/L/h) and yield (1.808mol/mol glucose) could in butanol to be competitive events. The productivity best fermentation performance of butanol was obtained in the casebe of obtained bagasse as Abstract the case of brick as immobilizing the butanol yield was 0.18 The highest hydrogen concentration of 66.76 % immobilizing carrier (5.804g/L of carrier, butanol while production, 0.22g/g of yield and g/g. 0.44g/L/h of productivity), while the hydrogen yield was obtained in the caseThe of activated carbonaddressed as productivity immobilizing carrier. just 1.41 mol/mol. highest hydrogen (1.808mol/mol glucose) could obtained in District heating networks are commonly in the(402mL/L/h) literature asand oneyield of the most effective solutions forbe decreasing the the case of brick as immobilizing carrier, while the butanol yield was 0.18 g/g. The highest hydrogen concentration of % greenhouse gas emissions frombytheElsevier building sector. These systems require high investments which are returned through66.76 the heat © 2019 The Authors. Published Ltd. was obtained in the case of activated carbon as immobilizing carrier. Copyright © 2018 Ltd. All rights reserved. sales. Due to theElsevier changed climate and building renovation policies, heat demand in the future could decrease, This is an open access article under theconditions CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/) Keywords: Co-production; Butanol; Hydrogen; Immobilization fermentation prolonging under the investment returnofperiod. Peer-review responsibility the scientific committee of ICAE2018 – The 10th International Conference on Applied Energy. Copyright 2018ofElsevier Ltd.isAll rights reserved. The main©scope this paper to assess the feasibility of using the heat demand – outdoor temperature function for heat demand Keywords: Co-production; Butanol; Hydrogen; Immobilization forecast. The district of Alvalade, located in Lisbon fermentation (Portugal), was used as a case study. The district is consisted of 665 1.buildings Introduction that vary in both construction period and typology. Three weather scenarios (low, medium, high) and three district renovation scenarios were developed (shallow, intermediate, deep). To estimate the error, obtained heat demand values were 1.compared Introduction from acondition dynamic heat previously developed of andnew validated by the Energy with is anresults essential for demand human model, survival, the exploration energy hasauthors. never been given up. The resultsthe showed that when changeofiswind, considered, the margin error could and be acceptable for some Nowadays technology foronly the weather exploitation hydrogen, solar,ofgeothermal nuclear energy is applications gradually Energy is annual an essential condition for survival, the exploration of new energy has never given up. (the error So in demand wasthe lower than 20% for all weather scenarios However, after introducing renovation maturing. researchers turn gaze tohuman the bioresource, hoping thatconsidered). it could reduce pollution and been simultaneously scenarios, the error value increased up to 59.5% (depending on the weather and renovation scenarios combination considered). Nowadays the technology for the exploitation of wind, hydrogen, solar, geothermal and nuclear energy is gradually gain more energy when the organic pollutants are used as the fermentation substrate. Among the various kinds of The valueSo ofbiobutanol slope coefficient increased on average within thehoping rangepromising of up toreduce 8% perpollution decade, that to the maturing. researchers turn the gazeare to recognized the bioresource, that3.8% it could andcorresponds simultaneously bioresource, and hydrogen as the most ones. decrease inenergy the number ofthe heating hours of 22-139h during the heating season (depending on Among the combination of weather and gain more when organic pollutants are used as the fermentation substrate. the various kinds of In contrast to ethanol, butanol can be blended with the gasoline. It can be used in existing motor engines and renovation scenarios considered). On theare other hand, function intercept increased ones. for 7.8-12.7% per decade (depending on the bioresource, biobutanol and hydrogen recognized as the most promising vehicular infrastructures without mechanical tailoring, and it canthebefunction transport with thefor existing pipelines for its low coupled scenarios). The values suggested could be used to the modify parameters the scenarios considered, and In contrast to.The ethanol, butanol canbutanol be blended with gasoline. It can bethat used in existing motor engines and vapor pressure energy value of (29.2 MJ/L) is 30% greater than of ethanol (21.2 MJ/L) and 10% improve the accuracy of heat demand estimations.
vehicular infrastructures without mechanical tailoring, and it can be transport with the existing pipelines for its low vapor pressure .The energy value of butanol (29.2 MJ/L) is 30% greater than that of ethanol (21.2 MJ/L) and 10% © 2017 The Authors. Published by Elsevier Ltd. Peer-review under responsibility of the Scientific Committee of The 15th International Symposium on District Heating and Cooling. * Corresponding author. Tel.:+86-028-85401057; fax:+86-028-85401057.
E-mail address: [email protected] Keywords: Heat demand; Forecast; Climate change * Corresponding author. Tel.:+86-028-85401057; fax:+86-028-85401057. E-mail address: [email protected] 1876-6102 Copyright © 2018 Elsevier Ltd. All rights reserved.
Selection and peer-review under responsibility of the scientific committee of the 10th International Conference on Applied Energy (ICAE2018). 1876-6102 Copyright © 2018 Elsevier Ltd. All rights reserved. Selection and peer-review responsibility of the scientific committee of the 10th International Conference on 1876-6102 © 2017 The Authors.under Published by Elsevier Ltd. Applied Energy (ICAE2018). Peer-review under responsibility of the Scientific Committee of The 15th International Symposium on District Heating and Cooling.
1876-6102 © 2019 The Authors. Published by Elsevier Ltd. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/) Peer-review under responsibility of the scientific committee of ICAE2018 – The 10th International Conference on Applied Energy. 10.1016/j.egypro.2019.01.435
1880 2
Jingyun Liu et al. / Energy Procedia 158 (2019) 1879–1884 Jingyun Liu/ Energy Procedia 00 (2018) 000–000
lesser than that of gasoline (32.5 MJ/L). Compared with ethanol, butanol is nonsensitivity to water, and less corrosiveness, lower volatility, less flammability and reduced toxicity to physical exposure [1]. Biohydrogen is known as an ideal renewable energy because of its high energy content (140MJ/kg) and lack of carbon emissions from combustion [2]. Acetone-butanol-ethanol (ABE) fermentation of clostridia is well known for butanol production from glucose and has been extensively investigated [3]. During ABE fermentation by clostridia, hydrogen is also produced. According to the current studies, butanol and hydrogen production are strongly affected by strains and culture. A butanol concentration of 13.1g/L, and a hydrogen production of 2.93L/L by Clostridium acetobutylicum ATCC824 have been got [4]. While in the study [5], butanol concentration and H2 production were 2.9g/L and 4.352L/L, respectively by Cl.spp fermentation. During batch fermentation for coproduction of butanol and hydrogen with suspend cell, biobutanol and biohydrogen production have some limitations, including lower yield, lower productivity, and lower final product concentration, lower utilization of the equipments. Fermentation in Continuous Stirring Tank Reactor (CSTR) can improve the utilization of the equipments. It was observed that short Hydraulic Retention Time (HRT) conditions in CSTR resulted in rapid decrease in biomass concentration, leading to decrease in volumetric hydrogen production. To overcome some of these, ABE production using immobilized cell systems with various Clostridium species have been studied with the hope of enhancing butanol productivity by achieving a high cell density. Many researchers had studied deeply in immobilizing cells, and most commonly employed immobilization methods are self-granulation, gel entrapment, surface attachment or biofilm systems. When it comes to the Cl.spp immobilization, surface attachment and gel entrapment are usually considered to be the popular two methods. In this study, surface attachment is chosen to immobilize cells, since the nutrients can directly contact with cells, with lower mass transfer resistance. The existing studies commonly have an eye on butanol or hydrogen individually. Few researchers have mentioned the co-production of butanol and hydrogen in immobilization fermentation. In the present study, activated carbon, bagasse and brick are used as the support materials to study the co-production performance of butanol and hydrogen. 2. Materials and method 2.1. Microorganisms and medium Clostridium acetobutylicum strain CICC8012 was used in the study, purchasing from China Center of Industrial Culture Collection (Beijing, China) in the form of freeze-dried powder. Approximately three rounds of pure cultivation later, the strain was suitable for fermentation. The medium of the seed culture and the fermentation contained: glucose 30g/L,yeast extraction 3g/L, phosphate buffer (KH2PO4 0.5g/L, K2HPO4 0.5g/L, ammonium acetate 2.2g/L), vitamins (para-amino-benzoic acid 1mg/L, thiamin 1mg/L, biotin 0.01mg/L), mineral salts (MgSO4·7H2O 0.2g/L, MnSO4·H2O 0.01g/L, FeSO4·7H2O 0.01g/L, NaCl 0.01g/L). Glucose and yeast extraction were sterilized by autoclaving at 121℃ for 20 min. Phosphate buffer, vitamins and mineral salts were filtersterilized through 0.45μ m sterile membrane. 2.2. The selection and processing of materials Activated carbon, bagasse and brick were the support materials in the experiment. Activated carbon was purchased from a supplier(Chengdu Cybernaut, China), the characteristical size of it was between 3-4mm. Bagasse was obtained from a local fruit stall and was cut into small and uniform size below 10mm. Brick was picked up from construction waste, then smashed and screened the particles which were 2-4mm. All of the materials were washed by running water and deionized water, then sterilized at 121℃ for 20minutes. The concentration of activated carbon and brick were 67g/L, but as for bagasse it was just 12.5g/L. 2.3. The procedure of the experiment Two rounds of experiment were conducted successively. The suspended cell fermentation and activated carbon immobilization fermentation were conducted in the first round. The remaining two materials as the immobilizing
Jingyun Liu et al. / Energy Procedia 158 (2019) 1879–1884 Jingyun Liu / Energy Procedia 00 (2018) 000–000
1881 3
material for fermentation were in the second round. There were totally forty conical flasks applied under every fermentation condition. Four of them were 250mL with the addition of 150mL medium (Two were used to collect the gas mixture and the other two were specially used to count the gas production by drainage method). The gas sample was collected with a gas sampling bag. The rest conical flasks were 50mL, which were filled with 40mL medium and they were capped with a tampon rather than rubber stopper for the release of gases to avoid the elevation of pressure internal. After inoculation with a ratio of 10% (V/V), conical flasks were incubated at 37℃ in a water bath. Two 50 ml conical flasks were picked out every four hours to determined glucose concentration, cell density and butanol concentration. At the same time, the gas contents in the sampling bag and drained water were also measured and weighed. 2.4. Analytical methods The glucose concentration in the broth was detected with DNS methods. Cell density in the suspended broth was determined with the spectrophotometer. Cells attached on the carrier materials were dried in the drying oven until constant weight and cell density was acquired by the difference of the materials’ weight before and after the experiment. The proportion of the hydrogen produced in the fermentation was analyzed with the a gas chromatograph (GC9790II, Zhejiang Fuli, China) equipped with a thermal conductivity detector and a 2m stainless column TDX-01. The carrier gas was N2 with the flow rate was 24ml/min. The temperature of the column oven was set at 90 °C, and the temperature was set at 100 °C for both the injection port and the detector. In the measurement of CO2, the hydrogen was chosen as the carrier gas with the flow rate of 40ml/min and the other settings were the same as hydrogen measurement. The concentration of acetone, butane and ethanol were also analyzed with a gas chromatograph equipped a flame ionization detector and a 30 m FFAP capillary column (i.d.0.32 mm), with N2 as the carrier gas. The oven temperature was increased from 60℃ to 180℃ at a rate of 20℃/min using a temperature program. The temperatures of the injector and the detector were set at 180 ℃ and 210℃ separately. Before the experiment, the glucose concentration and the cell density after inoculation were tested with the above analytical methods. 3. Results and discussion 3.1. Fermentation performances with different support materials as carrier The fermentation performances with activated carbon immobilization, bagasse and brick immobilization as well as suspended cell fermentation were illustrated in Fig.1. It could be seen that during the whole fermentation period, the butanol concentration, productivity and yield (5.804g/L ， 0.44g/L/h and 0.22g/g glucose) from bagasse immobilization fermentation were highest. The worst butanol production was obtained during fermentation by activated carbon immobilization. The detailed results were showed in Table 1. Different from butanol, hydrogen production is more directly related to glucose consumption and biomass growth. This phenomenon was appeared more clearly during the fermentation by suspended cell, bagasse immobilization and brick immobilization. When the fermentation finished, the highest hydrogen productivity 402mL/L/h and best yield 1.81mol/mol glucose were obtained during brick immobilization fermentation. But a high hydrogen proportion of 66.67% and a production 6.55L/L broth were obtained during activated carbon immobilization fermentation. Obviously, immobilization was indeed good for fermentation and different carrier materials had different effect on the cells. Compared with fermentation by suspend cell, the glucose consumption rate was increased 10.7%, 21.6%, 27.2% respectively by activated carbon, bagasse and brick immobilization. If bagasse was used as the support materials, butanol production was promoted. Activated carbon and brick immobilization were beneficial to hydrogen production.
Jingyun Liu et al. / Energy Procedia 158 (2019) 1879–1884 Jingyun Liu/ Energy Procedia 00 (2018) 000–000
2
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Fig. 1 Time course profiles of immobilization fermentation and suspended fermentation. (a) Suspended cell ; (b) Activated carbon immobilization; (c) Bagasse immobilization; (d) Brick immobilization
3.2. Co-production performance of butanol and hydrogen In the metabolic pathway of Clostridium acetobutylicum, there are two phases, acidogenesis and solventogenesis. The butyric acid and acetic acid are produced in the first phase along with glucose consumption, cell growth and hydrogen production. When the cell enters the stationary growth phase, the metabolism of the cells undergoes a shift to solvent production. From Fig. 1, butanol production was probably 8h later than hydrogen production, during the four kinds of fermentations. The conversion performance of glucose was illustrated in Fig.2. It could be found that enhanced fermentation was achieved if the cell was immobilized. The highest hydrogen yield could be achieved during fermentation by brick immobilization and the highest butanol yield could be achieved during fermentation by bagasse immobilization. Butanol can be absorbed in activated carbon, leading to higher butanol concentration for cell growth attached on activated carbon. From Fig.2 it could be also seen that competition events were existed between hydrogen and butanol. Compared with fermentation by bagasse immobilization, more hydrogen could be obtained but less butanol was produced during fermentation by brick immobilization.
Jingyun Liu et al. / Energy Procedia 158 (2019) 1879–1884 Jingyun Liu / Energy Procedia 00 (2018) 000–000 2.0
hydrogen yield butanol yield 1.44 1.15
1.2
1.42
0.25
1.81 0.19
0.15
0.15 0.1
0.8
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0.0
0.20
Butanol yield
Hydrogen yield
1.6
0.22
1883 5
0.05
suspended activated carbon
bagasse
brick
0.00
Fig. 2 Comparision of the hydrogen yield and butanol yield under different experiment conditions.
3.3. The reason for the enhancement from immobilization fermentation The change of metabolic pathway is recognized as the prime reason for the change of metabolites. In the metabolic process, the activity of enzyme is a decisive factor. The nature of enzyme is protein, which is coded by the relevant genes. It was reported that in the pathway of Clostridium acetobutylicum, the enzymes and component related with hydrogen and butanol production are ferredoxin, hydrogenase, NADH-ferredoxin oxidoreductase and so on [6]. Among them ferredoxin is an iron-sulfur-containing protein and under appropriate conditions the reduced ferredoxin is able to transfer electrons to an iron-containing hydrogenase which result in the production of molecular hydrogen. NADH-ferredoxin oxidoreductase is used in the electron transferring reaction, such as in the formation of hydrogen. Many of the genes for coding the above proteins are up-regulated and most of the genes involved in glycolysis, e.g., glcG, glcK, fba, gapN, gapC, pgk, tpi, pgm, eno, and pykA, which were probably related to the NADH-ferredoxin oxidoreductase were up-regulated in the immobilized cells at early fermentation phases. The induced expression of the hydGEFF and hydA2 at later fermentation phases was likely responsible for the increased hydrogen formation [7]. Two genes that are responsible for Fe2+uptake (CA_C1029 and CA_C1030) exhibited a 2 to 3 folds up-regulation throughout the fermentation. And the genes about the Fe 3+ uptake (CA_C0788-CA_C0791) were up-regulated 3 to 7 folds and sulfur uptake genes was also up-regulate. The regulation of the two genes was thought to form Fe–S clusters, which was involved in ferredoxin and hydrogenase. The competition events could be explained according to metabolic pathway. The electrons derived from the phosphoroclastic cleavage of pyruvate were transferred into hydrogen in the acid-producing metabolism. In the solvent-producing phase, electrons were transferred into NADPH, which was only emerged in the butanol production. In this study, different materials have different effect on the cells. The change in the metabolism was different and resulted in different outcomes. Table 1 The results of hydrogen and butanol Hydrogen Support Production Yield material L/L mol/mol Suspended Activated carbon Bagasse Brick
Productivity mL/L/h
Proportion
Concentration g/L
Butanol Yield g/g
Productivity g/L/h
5.25
1.15
301
61.64%
5.659
0.15
0.25
6.55
1.44
180
66.76%
3.832
0.10
0.43
4.77 6.07
1.42 1.81
209 402
51.58% 62.42%
5.804 4.922
0.22 0.19
0.44 0.28
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Jingyun Liu et al. / Energy Procedia 158 (2019) 1879–1884 Jingyun Liu/ Energy Procedia 00 (2018) 000–000
4. Conclusions Enhanced fermentation performance could be obtained by cell immobilization for co-production of hydrogen and butanol. Competition events were existed between hydrogen and butanol. The highest hydrogen yield could be achieved during fermentation by brick immobilization and the highest butanol yield could be achieved during fermentation by bagasse immobilization. Acknowledgements The present work was supported by the Fundamental Research Funds for the Central Universities (No.20822041B4013). References [1] Nanda S, Golemi-Kotra D, McDermott JC, Dalai AK, Gokalp I, Kozinski JA. Fermentative production of butanol: Perspectives on synthetic biology. New Biotechnol. 2017;37:210-21. [2] Gonzales RR, Sivagurunathan P, Parthiban A, Kim SH. Optimization of substrate concentration of dilute acid hydrolyzate of lignocellulosic biomass in batch hydrogen production. Int Biodeter Biodegr. 2016;113:22-7. [3] Lertsriwong S, Comwien J, Chulalaksananukul W, Glinwong C. Isolation and and identification of anaerobic bacteria from coconut wastewater factory for ethanol, butanol and 2, 3 butanediol production. Int Biodeter Biodegr. 2017;119:461-6. [4] Wang Y, Guo WQ, Cheng CL, Ho SH, Chang JS, Ren NQ. Enhancing bio-butanol production from biomass of Chlorella vulgaris JSC-6 with sequential alkali pretreatment and acid hydrolysis. Bioresource Technol. 2016;200:557-64. [5] Cheng CL, Che PY, Chen BY, Lee WJ, Lin CY, Chang JS. Biobutanol production from agricultural waste by an acclimated mixed bacterial microflora. Appl Energ. 2012;100:3-9. [6] Jones DT, Woods DR. Acetone-Butanol Fermentation Revisited. Microbiol Rev. 1986;50:484-524. [7] Liu D, Xu JH, Wang YY, Chen Y, Shen XN, Niu HQ, et al. Comparative transcriptomic analysis of Clostridium acetobutylicum biofilm and planktonic cells. Journal Of Biotechnology. 2016;218:1-12.
ScienceDirect Availableonline onlineatatwww.sciencedirect.com www.sciencedirect.com Available ScienceDirect Energy Procedia 00 (2018) 000–000 ScienceDirect Energy Procedia 00 (2018) 000–000 ScienceDirect
Available online at www.sciencedirect.com
Energy Procedia 158 Energy Procedia 00(2019) (2017)1879–1884 000–000
www.elsevier.com/locate/procedia www.elsevier.com/locate/procedia www.elsevier.com/locate/procedia
10th International Conference on Applied Energy (ICAE2018), 22-25 August 2018, Hong Kong, China 10th International Conference on Applied Energy (ICAE2018), 22-25 August 2018, Hong Kong, China StudyTheof15th theInternational co-production of on butanol and hydrogen Symposium District Heating and Coolingby
immobilizing Clostridium of acetobutylicum CICC8012 Study of the co-production butanol and hydrogen by * Assessing the feasibility of using the heat demand-outdoor immobilizing Clostridium Jingyun Liu, Wencan Zhou,acetobutylicum Senqing Fan , ZeyiCICC8012 Xiao * heat temperatureJingyun function for a long-term district demand forecast SchoolLiu, of Chemical Engineering, Sichuan University, Chengdu, 610065 Xiao Wencan Zhou, Senqing Fan , Zeyi
Abstract a
a a Sichuan University, c School of Chemical Engineering, 610065 I. Andrića,b,c*, A. Pina , P. Ferrão , J. Fournierb., Chengdu, B. Lacarrière , O. Le Correc
IN+ Center for Innovation, Technology and Policy Research - Instituto Superior Técnico, Av. Rovisco Pais 1, 1049-001 Lisbon, Portugal
b Abstract Three kinds of carrier materials, activated&carbon, bagasse and brick wereDaniel, used as immobilizing carriers during fermentation Veolia Recherche Innovation, 291 Avenue Dreyfous 78520 Limay, France c acetobutylicum CICC8012. Compared with cell suspended fermentation, enhanced fermentation performance was by Clostridium Département Systèmes Énergétiques et Environnement - IMT Atlantique, 4 rue Alfred Kastler, 44300 Nantes, France achieved cell fermentation, with bagasse shorter fermentation time required. During thecarriers experiments, and Three during kinds ofimmobilizing carrier materials, activated carbon, and brick were used as immobilizing duringhydrogen fermentation butanol appear acetobutylicum to be competitive events. The best fermentation performance of butanolenhanced was obtained in the case of bagassewas as by Clostridium CICC8012. Compared with cell suspended fermentation, fermentation performance immobilizing carrier (5.804g/Lcell of butanol production, 0.22g/gfermentation of yield andtime 0.44g/L/h of productivity), while the hydrogen achieved during immobilizing fermentation, with shorter required. During the experiments, hydrogenyield and was justappear 1.41 mol/mol. The highest hydrogen (402mL/L/h) and yield (1.808mol/mol glucose) could in butanol to be competitive events. The productivity best fermentation performance of butanol was obtained in the casebe of obtained bagasse as Abstract the case of brick as immobilizing the butanol yield was 0.18 The highest hydrogen concentration of 66.76 % immobilizing carrier (5.804g/L of carrier, butanol while production, 0.22g/g of yield and g/g. 0.44g/L/h of productivity), while the hydrogen yield was obtained in the caseThe of activated carbonaddressed as productivity immobilizing carrier. just 1.41 mol/mol. highest hydrogen (1.808mol/mol glucose) could obtained in District heating networks are commonly in the(402mL/L/h) literature asand oneyield of the most effective solutions forbe decreasing the the case of brick as immobilizing carrier, while the butanol yield was 0.18 g/g. The highest hydrogen concentration of % greenhouse gas emissions frombytheElsevier building sector. These systems require high investments which are returned through66.76 the heat © 2019 The Authors. Published Ltd. was obtained in the case of activated carbon as immobilizing carrier. Copyright © 2018 Ltd. All rights reserved. sales. Due to theElsevier changed climate and building renovation policies, heat demand in the future could decrease, This is an open access article under theconditions CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/) Keywords: Co-production; Butanol; Hydrogen; Immobilization fermentation prolonging under the investment returnofperiod. Peer-review responsibility the scientific committee of ICAE2018 – The 10th International Conference on Applied Energy. Copyright 2018ofElsevier Ltd.isAll rights reserved. The main©scope this paper to assess the feasibility of using the heat demand – outdoor temperature function for heat demand Keywords: Co-production; Butanol; Hydrogen; Immobilization forecast. The district of Alvalade, located in Lisbon fermentation (Portugal), was used as a case study. The district is consisted of 665 1.buildings Introduction that vary in both construction period and typology. Three weather scenarios (low, medium, high) and three district renovation scenarios were developed (shallow, intermediate, deep). To estimate the error, obtained heat demand values were 1.compared Introduction from acondition dynamic heat previously developed of andnew validated by the Energy with is anresults essential for demand human model, survival, the exploration energy hasauthors. never been given up. The resultsthe showed that when changeofiswind, considered, the margin error could and be acceptable for some Nowadays technology foronly the weather exploitation hydrogen, solar,ofgeothermal nuclear energy is applications gradually Energy is annual an essential condition for survival, the exploration of new energy has never given up. (the error So in demand wasthe lower than 20% for all weather scenarios However, after introducing renovation maturing. researchers turn gaze tohuman the bioresource, hoping thatconsidered). it could reduce pollution and been simultaneously scenarios, the error value increased up to 59.5% (depending on the weather and renovation scenarios combination considered). Nowadays the technology for the exploitation of wind, hydrogen, solar, geothermal and nuclear energy is gradually gain more energy when the organic pollutants are used as the fermentation substrate. Among the various kinds of The valueSo ofbiobutanol slope coefficient increased on average within thehoping rangepromising of up toreduce 8% perpollution decade, that to the maturing. researchers turn the gazeare to recognized the bioresource, that3.8% it could andcorresponds simultaneously bioresource, and hydrogen as the most ones. decrease inenergy the number ofthe heating hours of 22-139h during the heating season (depending on Among the combination of weather and gain more when organic pollutants are used as the fermentation substrate. the various kinds of In contrast to ethanol, butanol can be blended with the gasoline. It can be used in existing motor engines and renovation scenarios considered). On theare other hand, function intercept increased ones. for 7.8-12.7% per decade (depending on the bioresource, biobutanol and hydrogen recognized as the most promising vehicular infrastructures without mechanical tailoring, and it canthebefunction transport with thefor existing pipelines for its low coupled scenarios). The values suggested could be used to the modify parameters the scenarios considered, and In contrast to.The ethanol, butanol canbutanol be blended with gasoline. It can bethat used in existing motor engines and vapor pressure energy value of (29.2 MJ/L) is 30% greater than of ethanol (21.2 MJ/L) and 10% improve the accuracy of heat demand estimations.
vehicular infrastructures without mechanical tailoring, and it can be transport with the existing pipelines for its low vapor pressure .The energy value of butanol (29.2 MJ/L) is 30% greater than that of ethanol (21.2 MJ/L) and 10% © 2017 The Authors. Published by Elsevier Ltd. Peer-review under responsibility of the Scientific Committee of The 15th International Symposium on District Heating and Cooling. * Corresponding author. Tel.:+86-028-85401057; fax:+86-028-85401057.
E-mail address: [email protected] Keywords: Heat demand; Forecast; Climate change * Corresponding author. Tel.:+86-028-85401057; fax:+86-028-85401057. E-mail address: [email protected] 1876-6102 Copyright © 2018 Elsevier Ltd. All rights reserved.
Selection and peer-review under responsibility of the scientific committee of the 10th International Conference on Applied Energy (ICAE2018). 1876-6102 Copyright © 2018 Elsevier Ltd. All rights reserved. Selection and peer-review responsibility of the scientific committee of the 10th International Conference on 1876-6102 © 2017 The Authors.under Published by Elsevier Ltd. Applied Energy (ICAE2018). Peer-review under responsibility of the Scientific Committee of The 15th International Symposium on District Heating and Cooling.
1876-6102 © 2019 The Authors. Published by Elsevier Ltd. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/) Peer-review under responsibility of the scientific committee of ICAE2018 – The 10th International Conference on Applied Energy. 10.1016/j.egypro.2019.01.435
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Jingyun Liu et al. / Energy Procedia 158 (2019) 1879–1884 Jingyun Liu/ Energy Procedia 00 (2018) 000–000
lesser than that of gasoline (32.5 MJ/L). Compared with ethanol, butanol is nonsensitivity to water, and less corrosiveness, lower volatility, less flammability and reduced toxicity to physical exposure [1]. Biohydrogen is known as an ideal renewable energy because of its high energy content (140MJ/kg) and lack of carbon emissions from combustion [2]. Acetone-butanol-ethanol (ABE) fermentation of clostridia is well known for butanol production from glucose and has been extensively investigated [3]. During ABE fermentation by clostridia, hydrogen is also produced. According to the current studies, butanol and hydrogen production are strongly affected by strains and culture. A butanol concentration of 13.1g/L, and a hydrogen production of 2.93L/L by Clostridium acetobutylicum ATCC824 have been got [4]. While in the study [5], butanol concentration and H2 production were 2.9g/L and 4.352L/L, respectively by Cl.spp fermentation. During batch fermentation for coproduction of butanol and hydrogen with suspend cell, biobutanol and biohydrogen production have some limitations, including lower yield, lower productivity, and lower final product concentration, lower utilization of the equipments. Fermentation in Continuous Stirring Tank Reactor (CSTR) can improve the utilization of the equipments. It was observed that short Hydraulic Retention Time (HRT) conditions in CSTR resulted in rapid decrease in biomass concentration, leading to decrease in volumetric hydrogen production. To overcome some of these, ABE production using immobilized cell systems with various Clostridium species have been studied with the hope of enhancing butanol productivity by achieving a high cell density. Many researchers had studied deeply in immobilizing cells, and most commonly employed immobilization methods are self-granulation, gel entrapment, surface attachment or biofilm systems. When it comes to the Cl.spp immobilization, surface attachment and gel entrapment are usually considered to be the popular two methods. In this study, surface attachment is chosen to immobilize cells, since the nutrients can directly contact with cells, with lower mass transfer resistance. The existing studies commonly have an eye on butanol or hydrogen individually. Few researchers have mentioned the co-production of butanol and hydrogen in immobilization fermentation. In the present study, activated carbon, bagasse and brick are used as the support materials to study the co-production performance of butanol and hydrogen. 2. Materials and method 2.1. Microorganisms and medium Clostridium acetobutylicum strain CICC8012 was used in the study, purchasing from China Center of Industrial Culture Collection (Beijing, China) in the form of freeze-dried powder. Approximately three rounds of pure cultivation later, the strain was suitable for fermentation. The medium of the seed culture and the fermentation contained: glucose 30g/L,yeast extraction 3g/L, phosphate buffer (KH2PO4 0.5g/L, K2HPO4 0.5g/L, ammonium acetate 2.2g/L), vitamins (para-amino-benzoic acid 1mg/L, thiamin 1mg/L, biotin 0.01mg/L), mineral salts (MgSO4·7H2O 0.2g/L, MnSO4·H2O 0.01g/L, FeSO4·7H2O 0.01g/L, NaCl 0.01g/L). Glucose and yeast extraction were sterilized by autoclaving at 121℃ for 20 min. Phosphate buffer, vitamins and mineral salts were filtersterilized through 0.45μ m sterile membrane. 2.2. The selection and processing of materials Activated carbon, bagasse and brick were the support materials in the experiment. Activated carbon was purchased from a supplier(Chengdu Cybernaut, China), the characteristical size of it was between 3-4mm. Bagasse was obtained from a local fruit stall and was cut into small and uniform size below 10mm. Brick was picked up from construction waste, then smashed and screened the particles which were 2-4mm. All of the materials were washed by running water and deionized water, then sterilized at 121℃ for 20minutes. The concentration of activated carbon and brick were 67g/L, but as for bagasse it was just 12.5g/L. 2.3. The procedure of the experiment Two rounds of experiment were conducted successively. The suspended cell fermentation and activated carbon immobilization fermentation were conducted in the first round. The remaining two materials as the immobilizing
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material for fermentation were in the second round. There were totally forty conical flasks applied under every fermentation condition. Four of them were 250mL with the addition of 150mL medium (Two were used to collect the gas mixture and the other two were specially used to count the gas production by drainage method). The gas sample was collected with a gas sampling bag. The rest conical flasks were 50mL, which were filled with 40mL medium and they were capped with a tampon rather than rubber stopper for the release of gases to avoid the elevation of pressure internal. After inoculation with a ratio of 10% (V/V), conical flasks were incubated at 37℃ in a water bath. Two 50 ml conical flasks were picked out every four hours to determined glucose concentration, cell density and butanol concentration. At the same time, the gas contents in the sampling bag and drained water were also measured and weighed. 2.4. Analytical methods The glucose concentration in the broth was detected with DNS methods. Cell density in the suspended broth was determined with the spectrophotometer. Cells attached on the carrier materials were dried in the drying oven until constant weight and cell density was acquired by the difference of the materials’ weight before and after the experiment. The proportion of the hydrogen produced in the fermentation was analyzed with the a gas chromatograph (GC9790II, Zhejiang Fuli, China) equipped with a thermal conductivity detector and a 2m stainless column TDX-01. The carrier gas was N2 with the flow rate was 24ml/min. The temperature of the column oven was set at 90 °C, and the temperature was set at 100 °C for both the injection port and the detector. In the measurement of CO2, the hydrogen was chosen as the carrier gas with the flow rate of 40ml/min and the other settings were the same as hydrogen measurement. The concentration of acetone, butane and ethanol were also analyzed with a gas chromatograph equipped a flame ionization detector and a 30 m FFAP capillary column (i.d.0.32 mm), with N2 as the carrier gas. The oven temperature was increased from 60℃ to 180℃ at a rate of 20℃/min using a temperature program. The temperatures of the injector and the detector were set at 180 ℃ and 210℃ separately. Before the experiment, the glucose concentration and the cell density after inoculation were tested with the above analytical methods. 3. Results and discussion 3.1. Fermentation performances with different support materials as carrier The fermentation performances with activated carbon immobilization, bagasse and brick immobilization as well as suspended cell fermentation were illustrated in Fig.1. It could be seen that during the whole fermentation period, the butanol concentration, productivity and yield (5.804g/L ， 0.44g/L/h and 0.22g/g glucose) from bagasse immobilization fermentation were highest. The worst butanol production was obtained during fermentation by activated carbon immobilization. The detailed results were showed in Table 1. Different from butanol, hydrogen production is more directly related to glucose consumption and biomass growth. This phenomenon was appeared more clearly during the fermentation by suspended cell, bagasse immobilization and brick immobilization. When the fermentation finished, the highest hydrogen productivity 402mL/L/h and best yield 1.81mol/mol glucose were obtained during brick immobilization fermentation. But a high hydrogen proportion of 66.67% and a production 6.55L/L broth were obtained during activated carbon immobilization fermentation. Obviously, immobilization was indeed good for fermentation and different carrier materials had different effect on the cells. Compared with fermentation by suspend cell, the glucose consumption rate was increased 10.7%, 21.6%, 27.2% respectively by activated carbon, bagasse and brick immobilization. If bagasse was used as the support materials, butanol production was promoted. Activated carbon and brick immobilization were beneficial to hydrogen production.
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Fig. 1 Time course profiles of immobilization fermentation and suspended fermentation. (a) Suspended cell ; (b) Activated carbon immobilization; (c) Bagasse immobilization; (d) Brick immobilization
3.2. Co-production performance of butanol and hydrogen In the metabolic pathway of Clostridium acetobutylicum, there are two phases, acidogenesis and solventogenesis. The butyric acid and acetic acid are produced in the first phase along with glucose consumption, cell growth and hydrogen production. When the cell enters the stationary growth phase, the metabolism of the cells undergoes a shift to solvent production. From Fig. 1, butanol production was probably 8h later than hydrogen production, during the four kinds of fermentations. The conversion performance of glucose was illustrated in Fig.2. It could be found that enhanced fermentation was achieved if the cell was immobilized. The highest hydrogen yield could be achieved during fermentation by brick immobilization and the highest butanol yield could be achieved during fermentation by bagasse immobilization. Butanol can be absorbed in activated carbon, leading to higher butanol concentration for cell growth attached on activated carbon. From Fig.2 it could be also seen that competition events were existed between hydrogen and butanol. Compared with fermentation by bagasse immobilization, more hydrogen could be obtained but less butanol was produced during fermentation by brick immobilization.
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hydrogen yield butanol yield 1.44 1.15
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Fig. 2 Comparision of the hydrogen yield and butanol yield under different experiment conditions.
3.3. The reason for the enhancement from immobilization fermentation The change of metabolic pathway is recognized as the prime reason for the change of metabolites. In the metabolic process, the activity of enzyme is a decisive factor. The nature of enzyme is protein, which is coded by the relevant genes. It was reported that in the pathway of Clostridium acetobutylicum, the enzymes and component related with hydrogen and butanol production are ferredoxin, hydrogenase, NADH-ferredoxin oxidoreductase and so on [6]. Among them ferredoxin is an iron-sulfur-containing protein and under appropriate conditions the reduced ferredoxin is able to transfer electrons to an iron-containing hydrogenase which result in the production of molecular hydrogen. NADH-ferredoxin oxidoreductase is used in the electron transferring reaction, such as in the formation of hydrogen. Many of the genes for coding the above proteins are up-regulated and most of the genes involved in glycolysis, e.g., glcG, glcK, fba, gapN, gapC, pgk, tpi, pgm, eno, and pykA, which were probably related to the NADH-ferredoxin oxidoreductase were up-regulated in the immobilized cells at early fermentation phases. The induced expression of the hydGEFF and hydA2 at later fermentation phases was likely responsible for the increased hydrogen formation [7]. Two genes that are responsible for Fe2+uptake (CA_C1029 and CA_C1030) exhibited a 2 to 3 folds up-regulation throughout the fermentation. And the genes about the Fe 3+ uptake (CA_C0788-CA_C0791) were up-regulated 3 to 7 folds and sulfur uptake genes was also up-regulate. The regulation of the two genes was thought to form Fe–S clusters, which was involved in ferredoxin and hydrogenase. The competition events could be explained according to metabolic pathway. The electrons derived from the phosphoroclastic cleavage of pyruvate were transferred into hydrogen in the acid-producing metabolism. In the solvent-producing phase, electrons were transferred into NADPH, which was only emerged in the butanol production. In this study, different materials have different effect on the cells. The change in the metabolism was different and resulted in different outcomes. Table 1 The results of hydrogen and butanol Hydrogen Support Production Yield material L/L mol/mol Suspended Activated carbon Bagasse Brick
Productivity mL/L/h
Proportion
Concentration g/L
Butanol Yield g/g
Productivity g/L/h
5.25
1.15
301
61.64%
5.659
0.15
0.25
6.55
1.44
180
66.76%
3.832
0.10
0.43
4.77 6.07
1.42 1.81
209 402
51.58% 62.42%
5.804 4.922
0.22 0.19
0.44 0.28
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4. Conclusions Enhanced fermentation performance could be obtained by cell immobilization for co-production of hydrogen and butanol. Competition events were existed between hydrogen and butanol. The highest hydrogen yield could be achieved during fermentation by brick immobilization and the highest butanol yield could be achieved during fermentation by bagasse immobilization. Acknowledgements The present work was supported by the Fundamental Research Funds for the Central Universities (No.20822041B4013). References [1] Nanda S, Golemi-Kotra D, McDermott JC, Dalai AK, Gokalp I, Kozinski JA. Fermentative production of butanol: Perspectives on synthetic biology. New Biotechnol. 2017;37:210-21. [2] Gonzales RR, Sivagurunathan P, Parthiban A, Kim SH. Optimization of substrate concentration of dilute acid hydrolyzate of lignocellulosic biomass in batch hydrogen production. Int Biodeter Biodegr. 2016;113:22-7. [3] Lertsriwong S, Comwien J, Chulalaksananukul W, Glinwong C. Isolation and and identification of anaerobic bacteria from coconut wastewater factory for ethanol, butanol and 2, 3 butanediol production. Int Biodeter Biodegr. 2017;119:461-6. [4] Wang Y, Guo WQ, Cheng CL, Ho SH, Chang JS, Ren NQ. Enhancing bio-butanol production from biomass of Chlorella vulgaris JSC-6 with sequential alkali pretreatment and acid hydrolysis. Bioresource Technol. 2016;200:557-64. [5] Cheng CL, Che PY, Chen BY, Lee WJ, Lin CY, Chang JS. Biobutanol production from agricultural waste by an acclimated mixed bacterial microflora. Appl Energ. 2012;100:3-9. [6] Jones DT, Woods DR. Acetone-Butanol Fermentation Revisited. Microbiol Rev. 1986;50:484-524. [7] Liu D, Xu JH, Wang YY, Chen Y, Shen XN, Niu HQ, et al. Comparative transcriptomic analysis of Clostridium acetobutylicum biofilm and planktonic cells. Journal Of Biotechnology. 2016;218:1-12.