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Tamarix species, a possible non-food feedstock for bioethanol production
Special Abstracts / Journal of Biotechnology 150S (2010) S1–S576
tiple sugars are essential for simultaneous saccharification and fermentation applications. In this work optimal process operation conditions are explored for two types of yeasts in order to achieve bioethanol production from olive oil solid wastes. Saccharomyces cerevisiae and Klyveromyces marxianus were chosen for the simultaneous fermentation and saccharification of the olive pomace. Methods: Yeast type, temperature of the fermentation and type and concentration of the substrate were varied in several fermentation experiments in order to find the suitable operation conditions for ethanol production from the olive pomace. Indicator variables of the performance of the fermentation were yeast growth rate, ethanol production and residual sugar concentration in the growth medium. Results: It was found that the growth ability of S. cerevisiae belong to the range of 26 ◦ C to 47 ◦ C. However ethanol production was strongly inhibited at temperatures higher than 30 ◦ C. On the other hand K. marxianus has shown able to grow up to 47 ◦ C. However ethanol production is found only up to 40 ◦ C. Olive pomace up to 200 g/l presents no inhibitory effects either in yeast growth of ethanol production. Discussion: K. marxianus shows good fermentation properties for bioethanol production from olive oil residues. It can grow at high temperature with high production rate of ethanol i.e. 14.5 g/l were obtain at laboratory scale. Besides it shows about the same growth rates in the presence of olive pomace, so no inhibitory compound for this yeast are present on these residues.
S159
are located in the anaerobic anode chamber, oxidizing some organic materials and releasing electrons to the anode and protons to the solution (which pass through the membrane towards the cathode). The cathode is sparged with air to provide oxygen for the reactions of electrons, protons and oxygen (forming water). The system is completed with a wire between the electrodes, where electric current can be measured. A two-chamber microbial fuel cell was designed and built in our laboratory where carbon fibres were used as electrodes and Nafion 0125 protonselective membrane was placed between the two cells. Anaerobic sludge from a local biogas plant was applied in the anode chamber to study the stability of the system and to measure the electric power generated on one hand, and to investigate the structure and features of the biofilm grown on the surface of the anode, on the other hand. Fig. 1. doi:10.1016/j.jbiotec.2010.08.412 [P-B.60] Tamarix species, a possible non-food feedstock for bioethanol production C. Calisti 1 , F. Canganella 1 , M. Ruzzi 1,∗ , R. Valentini 3
Moresi 2 , M.
Petruccioli 1 , M.
1
doi:10.1016/j.jbiotec.2010.08.411 [P-B.59]
DABAC, University of Tuscia, Italy DISTA, University of Tuscia, Italy 3 DISAFRI, University of Tuscia, Italy Keywords: Bioethanol; Tamarix spp.; Fermentation; Direct steaminjection 2
Microbial fuel cells using anaerobic sludge Balázs Vajda ∗ , Katalin Bélafi-Bakó, Nándor Nemestóthy University of Pannonia, Hungary Keywords: Bio-electrochemical device; Wastewater; Microorganism; Protonselective membrane; Electricity Microbial fuel cells (MFCs) are bio-electrochemical devices capable to generate power by the activity of certain microorganisms. The traditional MFCs consist of an anode and a cathode chambers, separated by a cation selective membrane. The microbes
Fig. 1. Scheme of our microbial fuel cell.
Within the present interest in the use of non-food agricultural raw materials for biofuel production, we are conducting a laboratory-scale study to assess the techno-economic feasibility of converting Tamarix spp. biomasses, cultured in desert area, into bio-ethanol via fermentation. To this end, four species of Tamarix biomasses (namely T. gallica, T. jordanis, T. aphylla var. erecta and T. aphylla) were chemically characterized. Owing to its highest content in cellulose with respect to that in hemicellulose and lignin and its lowest phenol compound level, T. jordanis was selected as target raw material and used to outline a hypothetical process flow sheet, thus directing our research activities to the following unit operations: steam-injection heating of the liquid-solid mixture, enzymatic hydrolysis of polysaccharidic fractions, ethanolic fermentation and ethanol recovery from exhausted fermentation media by membrane processing. Most of the current biomass research has been focused on batch-driven laboratory techniques that are unable to extract useful information to transfer the process to a pilot-scale stage. To this end, we have developed a novel laboratory-scale direct steaminjection apparatus to perform the following operations: i) direct steam injection heating of a liquid-solid mixture in presence or absence of acid or alkaline catalyst up to 40 bar and 250 ◦ C; ii) steam-explosion; iii) liquid-solid separation; and iv) steam condensation and sub-cooling. Potential thermophilic and hyperthermophilic microbial candidates for the production of thermostable saccharolytic enzymes were screened. The yeast strains selected for genetic improvement of ethanol production, i.e. Saccharomyces bayanus BO 123, Saccharomyces cerevisiae F15 and Pichia anomala AN 4/3, exhibited good ethanol yields on model media and tolerance to inhibitors present in lignocellulosic hydrolysates. Finally, it was predicted a theoretical enrichment of clarified exhausted media from 3 to
S160
Special Abstracts / Journal of Biotechnology 150S (2010) S1–S576
8% v/v ethanol using commercial reverse-osmosis spiral-wound membrane modules provided that ethanol rejection ranged from 0.5 to 0.9. doi:10.1016/j.jbiotec.2010.08.413 [P-B.61] Inhibitory effect of crude glycerol on ethanol production by Enterobacter aerogenes Sang Jun Lee 1,∗ , Sung Bong Kim 1 , Seong Woo Kang 1 , Sung Ok Han 1 , Chulhwan Park 2 , Seung Wook Kim 1 1
Korea university, Korea, Republic of Kwangwoon university, Korea, Republic of Keywords: Crude glycerol; Enterobacter aerogenes; Ethanol production; Inhibitory effect 2
Glycerol utilization has a significant role on biodiesel production since large amount of glycerol is obtained as a by-product. Therefore, the development of new methods to convert crude and waste glycerol into higher value products is an urgent need. Recently, the glycerol as feedstock has been used to produce various value added products such as 1,3-propanediol, dihydroxyacetone, ethanol etc. through the biological process. Waste glycerol, which is generated from biodiesel manufacturing process, contains various inhibitory compounds (NaCl, KCl, biodiesel residue, and unreacted oils) and pH range of waste glycerol is very broad (pH 3-11). In this study, ethanol production from pure glycerol using Enterobacter aerogenes (ATCC 29007, 13048, 35028) was evaluated in anaerobic culture condition. Inhibitory effects of osmotic pressure of glycerol, pH, and salt concentrations were investigated based on waste glycerol ingredients. The ingredients of waste glycerol were supplied from a biodiesel manufacturing company in Korea. Ethanol production was performed with pure glycerol concentration of 5 g/L ∼ 30 g/L to evaluate effects of substrate concentration and osmotic pressure. The consumed glycerol was 3 ∼ 5 g/L and conversion rate was more than 0.9 mol-ethanol/molglycerol after 24 h of cultivation. To evaluate inhibitory effects of salts (NaCl and KCl), experiments were performed under 0 g/L ∼ 20 g/L of only one of each salt. Inhibitory effects of salts were shown at the high salt concentration. And then, inhibitory effect of pH was performed in the pH range of 4 ∼ 10 and cell growth and ethanol production was higher at pH 5 ∼ 7 than the others. doi:10.1016/j.jbiotec.2010.08.414 [P-B.62] Ethanol production from lignocellulosic materials by S. cerevisiae and P. stipitis Hawon Lee 1 , Yong Hwan Kim 1 , Sang Jun Lee 2,∗ , Seung Wook Kim 2 , Tak-Hyun Kim 3 , Chulhwan Park 1 1
Kwangwoon University, Korea, Republic of Korea University, Korea, Republic of 3 Korea Atomic Energy Research Institute, Korea, Republic of Keywords: Ethanol production; Lignocellulosic materials; S. cerevisiae; P. stipitis 2
The hydrolysis process of lignocelllosic materials to convert polysaccharides to monosaccharides is required in lignocellulosic ethanol production. However, during hydrolysis process various toxic compounds, which can inhibit ethanol production, are also produced and released. We investigated the performance of Saccharomyces cerevisiae K35 and Pichia stipitis KCCM 12009 in
the synthetic medium including model compounds (acetic acid, furfural, 5-hydroxymethylfurfural (5-HMF), syringaldehyde, and pcoumaric acid) as well as in lignocellulosic hydrolysates (yellow poplar, waste wood, and rice hull hydrolysates). In the case of fermentation by S. cerevisiae K35 in synthetic medium, the cell growth was inhibited with the increase in the acetic acid concentration but the ethanol yield was over 96% of the theoretical yield. In the high concentration (over 3 g/L), furfural and 5-HMF significantly decreased cell growth and ethanol production rate but not ethanol yield. S. cerevisiae K35 did not grow and did not produce ethanol at 5 g/L syringaldehyde and p-coumaric acid. In the fermentation of hydrolysates containing a variety of toxic compounds by S. cerevisiae K35, there was no synergetic effect of multiple inhibitory compounds. S. cerevisiae K35 showed high resistance, while P. stipitis KCCM 12009 was sensitive to inhibitory compounds comparison to S. cerevisiae K35. As the concentration of inhibitory compounds increased, cell growth of and ethanol production by P. stipitis KCCM 12009 decreased. P. stipitis KCCM 12009 was especially inhibited in the high concentration (5 g/L) of furans and phenolics. In the case of hydrolysates except yellow poplar hydrolysates, the ethanol production of P. stipitis KCCM 12009 was similar to that of the reference culture containing no inhibitory compounds. Acknowledgements: The authors gratefully acknowledge the financial support provided by the Korea Ministry of Environment (Eco-Star Project). This research was also supported by a Research Grant from Kwangwoon University in 2010. doi:10.1016/j.jbiotec.2010.08.415 [P-B.63] Fluorescence alternative of gram staining as a tool for metabolic switch recognition in butanol fermentation M. Linhova ∗ , P. Patakova, J. Lipovsky, P. Fribert, M. Rychtera, K. Melzoch Institue of Chemical Technology Prague, Czech Republic Keywords: Acetone-butanol-ethanol fermentation; Clostridium; Flow cytometry; Gram staining 1-butanol, a possible bio-component of liquid fuels with more advantageous properties in comparison with ethanol, can be produced by various strains of the genera Clostridium at so-called acetone-butanol-ethanol (ABE) fermentation using a sacharidic substrate. A cultivation course is typically biphasic when growth of cells is connected with butyrate and acetate formations and solvents production is achieved at cells sporulation. Therefore, detailed knowledge of clostridia behaviour is necessary for fermentation control. Although, the process is nowadays studied very intensively a deep attention is devoted mainly to few solventogenic strains like C.acetobutylicum ATCC 824 or C.beijerinckii NCIMB 8052. A combination of dyes hexidium iodide and SYTO 13 together with fluorescent microscopy and flow cytometry was used for recognition of acidogenic and solventogenic metabolic phases using species C.pasteurianum NRRL B-598, C.acetobutylicum DSM 1731 and C.beijerinckii CCM 6218. Cells in acidogenic state, showing G+ response if stained according to Gram, fluoresced bright red while solventogenic cells, having G− response at traditional Gram staining, were of green-yellow colour. The method was used for cells labelling during cultivation using glucose cultivation medium. Labelling patterns were significantly diverse in case of different strains and corresponded to determined concentrations of liquid (acids and solvents) and gaseous (H2 and CO2 ) metabolites. doi:10.1016/j.jbiotec.2010.08.416
tiple sugars are essential for simultaneous saccharification and fermentation applications. In this work optimal process operation conditions are explored for two types of yeasts in order to achieve bioethanol production from olive oil solid wastes. Saccharomyces cerevisiae and Klyveromyces marxianus were chosen for the simultaneous fermentation and saccharification of the olive pomace. Methods: Yeast type, temperature of the fermentation and type and concentration of the substrate were varied in several fermentation experiments in order to find the suitable operation conditions for ethanol production from the olive pomace. Indicator variables of the performance of the fermentation were yeast growth rate, ethanol production and residual sugar concentration in the growth medium. Results: It was found that the growth ability of S. cerevisiae belong to the range of 26 ◦ C to 47 ◦ C. However ethanol production was strongly inhibited at temperatures higher than 30 ◦ C. On the other hand K. marxianus has shown able to grow up to 47 ◦ C. However ethanol production is found only up to 40 ◦ C. Olive pomace up to 200 g/l presents no inhibitory effects either in yeast growth of ethanol production. Discussion: K. marxianus shows good fermentation properties for bioethanol production from olive oil residues. It can grow at high temperature with high production rate of ethanol i.e. 14.5 g/l were obtain at laboratory scale. Besides it shows about the same growth rates in the presence of olive pomace, so no inhibitory compound for this yeast are present on these residues.
S159
are located in the anaerobic anode chamber, oxidizing some organic materials and releasing electrons to the anode and protons to the solution (which pass through the membrane towards the cathode). The cathode is sparged with air to provide oxygen for the reactions of electrons, protons and oxygen (forming water). The system is completed with a wire between the electrodes, where electric current can be measured. A two-chamber microbial fuel cell was designed and built in our laboratory where carbon fibres were used as electrodes and Nafion 0125 protonselective membrane was placed between the two cells. Anaerobic sludge from a local biogas plant was applied in the anode chamber to study the stability of the system and to measure the electric power generated on one hand, and to investigate the structure and features of the biofilm grown on the surface of the anode, on the other hand. Fig. 1. doi:10.1016/j.jbiotec.2010.08.412 [P-B.60] Tamarix species, a possible non-food feedstock for bioethanol production C. Calisti 1 , F. Canganella 1 , M. Ruzzi 1,∗ , R. Valentini 3
Moresi 2 , M.
Petruccioli 1 , M.
1
doi:10.1016/j.jbiotec.2010.08.411 [P-B.59]
DABAC, University of Tuscia, Italy DISTA, University of Tuscia, Italy 3 DISAFRI, University of Tuscia, Italy Keywords: Bioethanol; Tamarix spp.; Fermentation; Direct steaminjection 2
Microbial fuel cells using anaerobic sludge Balázs Vajda ∗ , Katalin Bélafi-Bakó, Nándor Nemestóthy University of Pannonia, Hungary Keywords: Bio-electrochemical device; Wastewater; Microorganism; Protonselective membrane; Electricity Microbial fuel cells (MFCs) are bio-electrochemical devices capable to generate power by the activity of certain microorganisms. The traditional MFCs consist of an anode and a cathode chambers, separated by a cation selective membrane. The microbes
Fig. 1. Scheme of our microbial fuel cell.
Within the present interest in the use of non-food agricultural raw materials for biofuel production, we are conducting a laboratory-scale study to assess the techno-economic feasibility of converting Tamarix spp. biomasses, cultured in desert area, into bio-ethanol via fermentation. To this end, four species of Tamarix biomasses (namely T. gallica, T. jordanis, T. aphylla var. erecta and T. aphylla) were chemically characterized. Owing to its highest content in cellulose with respect to that in hemicellulose and lignin and its lowest phenol compound level, T. jordanis was selected as target raw material and used to outline a hypothetical process flow sheet, thus directing our research activities to the following unit operations: steam-injection heating of the liquid-solid mixture, enzymatic hydrolysis of polysaccharidic fractions, ethanolic fermentation and ethanol recovery from exhausted fermentation media by membrane processing. Most of the current biomass research has been focused on batch-driven laboratory techniques that are unable to extract useful information to transfer the process to a pilot-scale stage. To this end, we have developed a novel laboratory-scale direct steaminjection apparatus to perform the following operations: i) direct steam injection heating of a liquid-solid mixture in presence or absence of acid or alkaline catalyst up to 40 bar and 250 ◦ C; ii) steam-explosion; iii) liquid-solid separation; and iv) steam condensation and sub-cooling. Potential thermophilic and hyperthermophilic microbial candidates for the production of thermostable saccharolytic enzymes were screened. The yeast strains selected for genetic improvement of ethanol production, i.e. Saccharomyces bayanus BO 123, Saccharomyces cerevisiae F15 and Pichia anomala AN 4/3, exhibited good ethanol yields on model media and tolerance to inhibitors present in lignocellulosic hydrolysates. Finally, it was predicted a theoretical enrichment of clarified exhausted media from 3 to
S160
Special Abstracts / Journal of Biotechnology 150S (2010) S1–S576
8% v/v ethanol using commercial reverse-osmosis spiral-wound membrane modules provided that ethanol rejection ranged from 0.5 to 0.9. doi:10.1016/j.jbiotec.2010.08.413 [P-B.61] Inhibitory effect of crude glycerol on ethanol production by Enterobacter aerogenes Sang Jun Lee 1,∗ , Sung Bong Kim 1 , Seong Woo Kang 1 , Sung Ok Han 1 , Chulhwan Park 2 , Seung Wook Kim 1 1
Korea university, Korea, Republic of Kwangwoon university, Korea, Republic of Keywords: Crude glycerol; Enterobacter aerogenes; Ethanol production; Inhibitory effect 2
Glycerol utilization has a significant role on biodiesel production since large amount of glycerol is obtained as a by-product. Therefore, the development of new methods to convert crude and waste glycerol into higher value products is an urgent need. Recently, the glycerol as feedstock has been used to produce various value added products such as 1,3-propanediol, dihydroxyacetone, ethanol etc. through the biological process. Waste glycerol, which is generated from biodiesel manufacturing process, contains various inhibitory compounds (NaCl, KCl, biodiesel residue, and unreacted oils) and pH range of waste glycerol is very broad (pH 3-11). In this study, ethanol production from pure glycerol using Enterobacter aerogenes (ATCC 29007, 13048, 35028) was evaluated in anaerobic culture condition. Inhibitory effects of osmotic pressure of glycerol, pH, and salt concentrations were investigated based on waste glycerol ingredients. The ingredients of waste glycerol were supplied from a biodiesel manufacturing company in Korea. Ethanol production was performed with pure glycerol concentration of 5 g/L ∼ 30 g/L to evaluate effects of substrate concentration and osmotic pressure. The consumed glycerol was 3 ∼ 5 g/L and conversion rate was more than 0.9 mol-ethanol/molglycerol after 24 h of cultivation. To evaluate inhibitory effects of salts (NaCl and KCl), experiments were performed under 0 g/L ∼ 20 g/L of only one of each salt. Inhibitory effects of salts were shown at the high salt concentration. And then, inhibitory effect of pH was performed in the pH range of 4 ∼ 10 and cell growth and ethanol production was higher at pH 5 ∼ 7 than the others. doi:10.1016/j.jbiotec.2010.08.414 [P-B.62] Ethanol production from lignocellulosic materials by S. cerevisiae and P. stipitis Hawon Lee 1 , Yong Hwan Kim 1 , Sang Jun Lee 2,∗ , Seung Wook Kim 2 , Tak-Hyun Kim 3 , Chulhwan Park 1 1
Kwangwoon University, Korea, Republic of Korea University, Korea, Republic of 3 Korea Atomic Energy Research Institute, Korea, Republic of Keywords: Ethanol production; Lignocellulosic materials; S. cerevisiae; P. stipitis 2
The hydrolysis process of lignocelllosic materials to convert polysaccharides to monosaccharides is required in lignocellulosic ethanol production. However, during hydrolysis process various toxic compounds, which can inhibit ethanol production, are also produced and released. We investigated the performance of Saccharomyces cerevisiae K35 and Pichia stipitis KCCM 12009 in
the synthetic medium including model compounds (acetic acid, furfural, 5-hydroxymethylfurfural (5-HMF), syringaldehyde, and pcoumaric acid) as well as in lignocellulosic hydrolysates (yellow poplar, waste wood, and rice hull hydrolysates). In the case of fermentation by S. cerevisiae K35 in synthetic medium, the cell growth was inhibited with the increase in the acetic acid concentration but the ethanol yield was over 96% of the theoretical yield. In the high concentration (over 3 g/L), furfural and 5-HMF significantly decreased cell growth and ethanol production rate but not ethanol yield. S. cerevisiae K35 did not grow and did not produce ethanol at 5 g/L syringaldehyde and p-coumaric acid. In the fermentation of hydrolysates containing a variety of toxic compounds by S. cerevisiae K35, there was no synergetic effect of multiple inhibitory compounds. S. cerevisiae K35 showed high resistance, while P. stipitis KCCM 12009 was sensitive to inhibitory compounds comparison to S. cerevisiae K35. As the concentration of inhibitory compounds increased, cell growth of and ethanol production by P. stipitis KCCM 12009 decreased. P. stipitis KCCM 12009 was especially inhibited in the high concentration (5 g/L) of furans and phenolics. In the case of hydrolysates except yellow poplar hydrolysates, the ethanol production of P. stipitis KCCM 12009 was similar to that of the reference culture containing no inhibitory compounds. Acknowledgements: The authors gratefully acknowledge the financial support provided by the Korea Ministry of Environment (Eco-Star Project). This research was also supported by a Research Grant from Kwangwoon University in 2010. doi:10.1016/j.jbiotec.2010.08.415 [P-B.63] Fluorescence alternative of gram staining as a tool for metabolic switch recognition in butanol fermentation M. Linhova ∗ , P. Patakova, J. Lipovsky, P. Fribert, M. Rychtera, K. Melzoch Institue of Chemical Technology Prague, Czech Republic Keywords: Acetone-butanol-ethanol fermentation; Clostridium; Flow cytometry; Gram staining 1-butanol, a possible bio-component of liquid fuels with more advantageous properties in comparison with ethanol, can be produced by various strains of the genera Clostridium at so-called acetone-butanol-ethanol (ABE) fermentation using a sacharidic substrate. A cultivation course is typically biphasic when growth of cells is connected with butyrate and acetate formations and solvents production is achieved at cells sporulation. Therefore, detailed knowledge of clostridia behaviour is necessary for fermentation control. Although, the process is nowadays studied very intensively a deep attention is devoted mainly to few solventogenic strains like C.acetobutylicum ATCC 824 or C.beijerinckii NCIMB 8052. A combination of dyes hexidium iodide and SYTO 13 together with fluorescent microscopy and flow cytometry was used for recognition of acidogenic and solventogenic metabolic phases using species C.pasteurianum NRRL B-598, C.acetobutylicum DSM 1731 and C.beijerinckii CCM 6218. Cells in acidogenic state, showing G+ response if stained according to Gram, fluoresced bright red while solventogenic cells, having G− response at traditional Gram staining, were of green-yellow colour. The method was used for cells labelling during cultivation using glucose cultivation medium. Labelling patterns were significantly diverse in case of different strains and corresponded to determined concentrations of liquid (acids and solvents) and gaseous (H2 and CO2 ) metabolites. doi:10.1016/j.jbiotec.2010.08.416