- Email: [email protected]
Increased performance of continuous stirred tank reactor with calcium supplementation
international journal of hydrogen energy 35 (2010) 2622–2626
Available at www.sciencedirect.com
journal homepage: www.elsevier.com/locate/he
Increased performance of continuous stirred tank reactor with calcium supplementation Zhuliang Yuan, Haijun Yang, Xiaohua Zhi, Jianquan Shen* Beijing National Laboratory for Molecular Sciences (BNLMS), New Materials Laboratory, Institute of Chemistry, Chinese Academy of Sciences, Beijing 100190, PR China
article info
abstract
Article history:
Continuous biohydrogen production with calcium supplementation at low hydraulic
Received 8 April 2009
retention time (HRT) in a continuous stirred tank reactor (CSTR) was studied to maximize
Received in revised form
the hydrogen productivity of anaerobic mixed cultures. After stable operations at HRT of
12 April 2009
8–4 h, the bioreactor became unstable when the HRT was lowered to 2 h. Supplementation
Accepted 12 April 2009
of 100 mg/L calcium at HRT 2 h improved the operation stability through enhancement of
Available online 20 May 2009
cell retention with almost two-fold increase in cell density than that without calcium
Keywords:
mol sucrose, respectively, both of which were the highest values our group have ever
Biohydrogen production
achieved. The results showed that calcium supplementation can be an effective way to
Anaerobic mixed cultures
improve the performance of CSTR at low HRT.
addition. Hydrogen production rate and hydrogen yield reached 24.5 L/d/L and 3.74 mol H2/
Continuous stirred tank reactor
ª 2009 Professor T. Nejat Veziroglu. Published by Elsevier Ltd. All rights reserved.
Calcium
1.
Introduction
As for the increasing pressure of energy crisis and environmental problems derivate from exhausting of fossil fuels, much attention has been paid to explore new energy sources. Hydrogen is among the most promising alternatives because it is energy intensive, clean, and renewable [1]. Compared to electrochemical and thermochemical hydrogen production processes, biohydrogen production is more environmentally friendly and cost effective. Up to now, many researches have been performed on anaerobic fermentation owing to the fact that hydrogen can be generated continuously at a high rate from renewable organic materials or biomass wastes in the absence of light [2,3]. Compared to pure cultures, fermentative hydrogen production processes using mixed cultures take in various kinds of non-sterile feedstock and may be suitable for commercial development. Thus many research groups
devoted their efforts to develop novel mixed cultures and improve their hydrogen productivity [4–8]. As for continuous hydrogen production, it is of great importance to maintain a sufficient amount of hydrogenproducing bacterial population in the bioreactor. Most studies on fermentative hydrogen production using mixed cultures have been conducted in conventional CSTR under mesophilic conditions. However, it is relatively difficult to achieve that in a CSTR because washout of the biomass usually occurs at a low hydraulic retention time (HRT). Attempts to enhance biomass retention by physical or biological immobilization of cells were shown to attain better hydrogen production performance than that of conventional CSTR, with hydrogen production rates ranging from 6–360 L/d/L [9–11]. Nevertheless, the matrices used for cell immobilization inevitably occupy significant space in the reactor, limiting cell density and possibly generating mass transfer barriers between
* Corresponding author. Tel.: þ86 10 62620903; fax: þ86 10 62559373. E-mail address: [email protected] (J. Shen). 0360-3199/$ – see front matter ª 2009 Professor T. Nejat Veziroglu. Published by Elsevier Ltd. All rights reserved. doi:10.1016/j.ijhydene.2009.04.018
international journal of hydrogen energy 35 (2010) 2622–2626
substrates and products. To avoid these problems, granular sludge was generated to enhance cell retention and biomass concentration simultaneously [12]. Biofilm-based systems were also used for anaerobic hydrogen production since they were more capable of maintaining higher biomass concentration and could operate at low HRT without biomass washout problems [13]. Some metal ions, such as calcium ion, were supplemented as coagulant aids to enhance the biomass accumulation [14]. However, the feasibilities of using the granular sludge or biofilm systems rely on the ability to control the timing and conditions of sludge granulation or biofilm formation as well as to maintain the stability and activity of granular sludge or biofilm. Furthermore, it is usually time-consuming to start up an up-flow anaerobic sludge blanket (UASB) or biofilm-based reactor, and poor startup of biological hydrogen production systems cause an ineffective hydrogen production rate and poor biomass growth at a high HRT, or cause a prolonged period of acclimation. An attractive and effective way may be that increasing the cell density in the reactor without immobilization matrices, as well as to ensure a relatively short start-up time and make the reactor easy to operate. In the consideration above, a conventional CSTR was utilized to study the maximization of hydrogen productivity using the well-mixed cultures used in our previous studies [6], owing to that CSTR is simple, easy to operate and low-cost without immobilization carrier. Furthermore, it usually takes shorter start-up time compared with UASB and biofilm systems. In order to improve the operation stability and maintain a high biomass density at relatively low HRT aiming to obtain a high hydrogen production rate as well as a high substrate utilizing rate, 100 mg/L calcium ions were added into the feed medium. The effect of calcium supplementation on the performance of CSTR was investigated by examining the hydrogen characteristics and metabolites distributions all over the operation course.
2.
Experimental section
2.1.
Inoculum, medium and analytical procedures
The inoculum of mixed cultures with mainly Clostridium pasteurianum bacteria, the synthetic medium with 11.1 g/L sucrose and the analytical procedures were the same as our previous work described in Ref [6].
2.2.
Operating conditions
The CSTR used in this study was 1 L in working volume with an internal diameter of 8 cm and a liquid height of 20 cm. The reactor was operated at 35 C and stirred by gas circulation. The HRT reduction was conducted in a stepwise manner through 78 days, respectively. At each HRT, the reactor was operated for about 2 weeks to allow steady-state condition development. Steady-state conditions were established when the variation in the product concentrations was constant (gas production rate, 5%; substrate degradation efficiency, 5%). After the constant parameter data were obtained, HRT was then shortened. Calcium with concentration of 100 mg/L was
2623
added in the feed medium at both 4 h and 2 h HRTs for comparison.
3.
Results and discussions
3.1. Reactor operation and hydrogen production characteristics Compared to UASB systems, which usually need more than 30 days to start up [12], it is time saving to start up a CSTR system. At an HRT of 8 h, the reactor achieved constant digestion gas production after 10 days of acclimation. The reactor needed less than 2 days to achieve constant digestion gas production after HRT reduction at each step except for the HRT reduction from 4 h to 2 h without calcium supplementation. The reactor was monitored by examining the effluent wastewater every three days for substrate degradation efficiency (Sd), hydrogen content (HC), hydrogen production rate (HPR), hydrogen yield (HY), volatile suspended solids (VSS), total suspended solids (TSS) and metabolites concentrations. As shown in Fig 1, the CSTR operated in a smooth state at HRTs of 8 h and 6 h. At both HRTs, the sucrose degradation efficiencies were over 98%. Gas product analyses indicated that the biogas only consisted of hydrogen and carbon dioxide, and the HCs were constantly 43% and 50% at HRT 8 h and HRT 6 h, respectively. The HPRs were 6.1 L/d/L and 10.0 L/d/L at HRT 8 h and HRT 6 h, respectively. From the VSS data we could see that the total amount of bacteria increased along with the decrease of the HRT, resulting in the increase of the hydrogen-producing bacteria and hydrogen production rate. When HRT decreased from 6 h to 4 h, sucrose degradation efficiency decreased to 89%, which was mainly due to the cell washout effect, because VSS value decreased from1054 at HRT 6 h to 889 mg/L at HRT 4 h caused by the increase of the substrate loading rate. Fortunately, the HC, HPR and HY all increased to the maxima of 55%, 14 L/d/L and 3.63 mol H2/mol sucrose, respectively. The reason for this phenomenon was probably that the content of hydrogen-producing bacteria in the mixed cultures increased along with the decrease of HRT, in spite of the decrease of total bacterial population. It was reported that hydrogen-producing bacteria reproduced much faster than other bacteria, so at relatively lower HRT, their content in the bacterial community usually increases with the decrease of HRT [15]. The reactor became quite unstable when HRT decreased to 2 h. All the parameters fluctuated largely during 15-day operation. Both the sucrose degradation efficiency and hydrogen content decreased to about 39%, which were the lowest during the whole operation course. HPR decreased to about 12.8 L/d/L, which was lower than that of 4 h HRT. Nevertheless, the hydrogen yield increased a little to 3.76 mol H2/mol sucrose, and was relatively constant despite of the fluctuations of Sd and HY. This was probably because that the ratio of the hydrogen production bacteria in the community was constant although the total cell density of the whole bacteria community changed with the environmental parameters [16]. In order to achieve a stable operation bioreactor at low HRT aiming to maximize the hydrogen productivity of hydrogen-producing bacteria, 100 mg/L calcium ions
2624
international journal of hydrogen energy 35 (2010) 2622–2626
Fig. 1 – Performance of CSTR: (a) hydrogen retention time (HRT), (b) substrate degradation efficiency (Sd) and hydrogen content (HC), (c) hydrogen production rate (HPR) and hydrogen yield (HY), (d) volatile suspended solids (VSS) and metabolites concentrations.
were added into the feed medium at HRT 2 h. When calcium was added into the medium at 2 h HRT, the substrate degradation efficiency increased to 73%, which was almost twice of that without calcium supplementation. HPR and HY increased to 24.5 L/d/L and 3.74 mol H2/mol sucrose, which were the highest values our research group has achieved and were relatively high compared with other hydrogen-producing bacteria in the CSTR mode [17,18]. Table 1 shows that VSS increased to 1682 mg/L, which was much higher than 747 mg/ L than that without calcium supplementation, indicating that cell retention in the reactor was the main reason for this improvement. Another reason was probably that calcium was required for catalytic activity, either as a participant in the reaction, or as a structural requirement in order to maintain the appropriate confirmation of the active site [12].
Interestingly, compared to UASB or biofilm-based systems with calcium supplementation [12,13], bacteria in our CSTR were always maintained in a suspended state and no granular sludge or biofilm formed during the whole operation, indicating an alternative way to improve performance of bioreactor by merely increasing the cell density, as well as avoiding the disadvantages of UASB or biofilm-based systems. As for comparison, with calcium supplementation at HRT 4 h, the sucrose degradation efficiency increased highly to 99%, and all the operation parameters almost remained constant during 15-day operation, indicating that the bioreactor became quite more stable mainly due to cell retention exhibiting the increase of VSS from 889 to 1092 mg/L. However, the HPR and HY decreased to about 11.2 L/d/L and 2.58 mol H2/mol sucrose, respectively. This might be that the
Table 1 – Substrate degradation efficiencies, biomass and hydrogen production characteristics. HRT (h) 8 6 4 2 4 2 a b c d e
Ca2þ (mg/L) 0 0 0 0 100 100
Sda (%)
HCb (%)
HPRc (L/d/L)
HYd (mol H2/mol sucrose)
99 1 99 1 89 4 38 9 99 73 5
43 50 55 39 3 42 1 47 2
6.1 0.1 10.0 0.2 14.0 1.3 12.8 2.5 11.2 24.5 0.5
2.80 0.01 3.48 0.03 3.63 0.12 3.76 0.22 2.58 3.74 0.32
Substrate degradation efficiency. Hydrogen content. Hydrogen production rate. Hydrogen yield. Ratio of volatile suspended solids and total suspended solids.
VSS (mg/L)
VSS/TSSe (%)
874 127 1054 80 889 152 747 167 1092 82 1682 470
87 93 86 90 73 72
2625
international journal of hydrogen energy 35 (2010) 2622–2626
Table 2 – pH values and metabolites distributions. HRT (h) 8 6 4 2 4 2 a b c d e f
Ca2þ (mg/L)
pH
0 0 0 0 100 100
5.70 0.05 5.40 0.19 5.44 0.12 6.47 0.11 5.16 0.27 5.35 0.26
EtOHa (mg/L) 760 80 610 10 510 120 1280 390 300 300 30
HAcb (mg/L)
HPrc (mg/L)
HBud (mg/L)
(A þ B)e (mg/L)
A/Bf
800 30 1340 20 1750 220 2370 230 570 140 2710 400
640 30 470 110 830 170 600 370 190 60 300 180
1060 40 1240 20 1730 230 1500 330 2260 10 1280 260
1860 2580 3480 3870 2830 3990
0.75 1.08 1.01 1.58 0.25 2.12
Ethanol. Acetate. Propionate. Butyrate. Sum of acetate and butyrate. Acetate/butyrate ratio.
bacterial community changed when calcium was added into the medium.
3.2.
Metabolites distributions
As for C. pasteurianum, which was dominant in a CSTR fermentor seeded with this kind of suspended mixed cultures, a theoretical maximum of 4 mol H2 per mol of hexose is obtained when acetic acid is the resulting product. When butyrate is the sole resulting product, a theoretical maximum of 2 mol H2 per mol of hexose is obtained [19]. In practice, high H2 yields are usually associated with a mixture of acetate and butyrate as fermentation products, and low H2 yields are associated with propionate and reduced resulting products (alcohols, lactic acid) [20]. Table 2 shows that the major products were butyrate, acetate, ethanol and propionate with average concentrations of 1060–2260, 570–2710, 300–1280 and 190–830 mg/L, respectively. As the HRT decreased from 8 h to 2 h without calcium supplementation, the sum of butyrate and acetate increased from 1860 to 3870 mg/L, which coincided with the increasing trend of HY. With the decrease of HRT from 8 h to 2 h, the acetate/butyrate ratio also showed an increasing trend except at HRT 4 h, which supported that higher concentration of acetate was benefit to hydrogen production. The decrease of acetate/butyrate ratio at HRT 4 h did not lower the HY, showing that the sum of acetate and butyrate determined higher HY under this condition. At HRT 2 h with calcium supplementation, the sum of acetate and butyrate and the acetate/butyrate ratio both increased to 3990 mg/L and 2.12 respectively, which were the highest during the whole operation course, which were consistent with the highest HY of 3.74 mol H2/mol sucrose. It proved again the conclusion that calcium supplementation at lower HRT improves the hydrogen productivity and alters the fermentation pathway to produce more hydrogen.
4.
Conclusions
The maximization of hydrogen productivity of suspended hydrogen-producing mixed cultures in a conventional CSTR was studied with calcium supplementation at low HRT. Operations of CSTR at HRT of 8–4 h without calcium
supplementation were stable with substrate degradation efficiencies more than 89%. The HPR and HY reached the peak at 14 L/d/L and 3.63 mol H2/mol sucrose at HRT 4 h, respectively. However, when the bioreactor was operated at a low HRT of 2 h, operation of bioreactor became quite unstable. Thus 100 mg/L calcium ions were added into the feed medium of HRT 2 h in order to maintain a high cell density in the fermentor. The bioreactor became more stable and the substrate degradation efficiency increased to 73%, which was twice that of bioreactor without calcium supplementation. HPR and HY were increased dramatically to 24.5 L/d/L and 3.74 mol H2/mol sucrose, respectively, which were the highest values we have ever reached. Retaining a high cell density and optimization of bacterial community in the bioreactor will be equally important to achieve higher hydrogen productivity and a better performance.
Acknowledgments The authors would like to thank the Ministry of Science & Technology, China for financial support under the National Hi-Tech R&D Program (863 Program, Grant No. 2006AA 05Z101).
references
[1] Lubitz W, Tumas W. Hydrogen: an overview. Chem Rev 2007; 107:3900–3. [2] Hallenbeck PC, Benemann JR. Biological hydrogen production: fundamentals and limiting processes. Int J Hydrogen Energy 2002;27:1185–93. [3] Kotay SM, Das D. Biohydrogen as a renewable energy resource-Prospects and potentials. Int J Hydrogen Energy 2008;33:258–63. [4] Lin CY, Lay CH. Carbon/nitrogen-ratio effect on fermentative hydrogen production by mixed microflora. Int J Hydrogen Energy 2004;29:41–5. [5] Van Ginkel SW, Oh SE, Logan BE. Biohydrogen gas production from food processing and domestic wastewaters. Int J Hydrogen Energy 2005;30:1535–42. [6] Yuan Z, Yang H, Zhi X, Shen J. Enhancement effect of l-cysteine on dark fermentative hydrogen production. Int J Hydrogen Energy 2008;33:6535–40.
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international journal of hydrogen energy 35 (2010) 2622–2626
[7] Ren N, Li J, Li B, Wang Y, Liu S. Biohydrogen production from molasses by anaerobic fermentation with a pilotscale bioreactor system. Int J Hydrogen Energy 2006;31: 2147–57. [8] Yu HQ, Mu Y. Biological hydrogen production in a UASB reactor with granules. II: reactor performance in 3-year operation. Biotechnol Bioeng 2006;94:988–95. [9] Wu SY, Hung CH, Lin CN, Chen HW, Lee AS, Chang JS. Fermentative hydrogen production and bacterial community structure in high-rate anaerobic bioreactors containing silicone-immobilized and self-flocculated sludge. Biotechnol Bioeng 2006;93:934–46. [10] Wu SY, Lin CN, Chang JS. Hydrogen production with immobilized sewage sludge in three-phase fluidized-bed bioreactors. Biotechnol Prog 2003;19:828–32. [11] Lee KS, Lo YC, Lin PJ, Chang JS. Improving biohydrogen production in a carrier-induced granular sludge bed by altering physical configuration and agitation pattern of the bioreactor. Int J Hydrogen Energy 2006;31:1648–57. [12] Yu H, Zhu Z, Hu W, Zhang H. Hydrogen production from rice winery wastewater in an upflow anaerobic reactor by using mixed anaerobic cultures. Int J Hydrogen Energy 2002;27: 1359–65. [13] Ahn Y, Park EJ, Oh YK, Park S, Webster G, Weightman AJ. Biofilm microbial community of a thermophilic trickling
[14]
[15]
[16]
[17]
[18]
[19]
[20]
biofilter used for continuous biohydrogen production. FEMS Microbiol Lett 2005;249:31–8. Kim IS, Jang N. The effect of calcium on the membrane biofouling in the membrane bioreactor (MBR). Water Res 2006;40:2756–64. Sahlstrom L. A review of survival of pathogenic bacteria in organic waste used in biogas plants. Bioresour Technol 2003; 87:161–6. Chang FY, Lin CY. Biohydrogen production using an up-flow anaerobic sludge blanket reactor. Int J Hydrogen Energy 2004; 29:33–9. Gavala HN, Skiadas IV, Ahring BK. Biological hydrogen production in suspended and attached growth anaerobic reactor systems. Int J Hydrogen Energy 2006;31:1164–75. Kim SH, Han SK, Shin HS. Performance comparison of a continuous-flow stirred-tank reactor and an anaerobic sequencing batch reactor for fermentative hydrogen production depending substrate concentration. Water Sci Technol 2005;52:23–9. Kleerebezem R, Stams AJ. Kinetics of syntrophic cultures: a theoretical treatise on butyrate fermentation. Biotechnol Bioeng 2000;67:529–43. Vignais PM, Magnin JP, Willison JC. Increasing biohydrogen production by metabolic engineering. Int J Hydrogen Energy 2006;31:1478–83.
Available at www.sciencedirect.com
journal homepage: www.elsevier.com/locate/he
Increased performance of continuous stirred tank reactor with calcium supplementation Zhuliang Yuan, Haijun Yang, Xiaohua Zhi, Jianquan Shen* Beijing National Laboratory for Molecular Sciences (BNLMS), New Materials Laboratory, Institute of Chemistry, Chinese Academy of Sciences, Beijing 100190, PR China
article info
abstract
Article history:
Continuous biohydrogen production with calcium supplementation at low hydraulic
Received 8 April 2009
retention time (HRT) in a continuous stirred tank reactor (CSTR) was studied to maximize
Received in revised form
the hydrogen productivity of anaerobic mixed cultures. After stable operations at HRT of
12 April 2009
8–4 h, the bioreactor became unstable when the HRT was lowered to 2 h. Supplementation
Accepted 12 April 2009
of 100 mg/L calcium at HRT 2 h improved the operation stability through enhancement of
Available online 20 May 2009
cell retention with almost two-fold increase in cell density than that without calcium
Keywords:
mol sucrose, respectively, both of which were the highest values our group have ever
Biohydrogen production
achieved. The results showed that calcium supplementation can be an effective way to
Anaerobic mixed cultures
improve the performance of CSTR at low HRT.
addition. Hydrogen production rate and hydrogen yield reached 24.5 L/d/L and 3.74 mol H2/
Continuous stirred tank reactor
ª 2009 Professor T. Nejat Veziroglu. Published by Elsevier Ltd. All rights reserved.
Calcium
1.
Introduction
As for the increasing pressure of energy crisis and environmental problems derivate from exhausting of fossil fuels, much attention has been paid to explore new energy sources. Hydrogen is among the most promising alternatives because it is energy intensive, clean, and renewable [1]. Compared to electrochemical and thermochemical hydrogen production processes, biohydrogen production is more environmentally friendly and cost effective. Up to now, many researches have been performed on anaerobic fermentation owing to the fact that hydrogen can be generated continuously at a high rate from renewable organic materials or biomass wastes in the absence of light [2,3]. Compared to pure cultures, fermentative hydrogen production processes using mixed cultures take in various kinds of non-sterile feedstock and may be suitable for commercial development. Thus many research groups
devoted their efforts to develop novel mixed cultures and improve their hydrogen productivity [4–8]. As for continuous hydrogen production, it is of great importance to maintain a sufficient amount of hydrogenproducing bacterial population in the bioreactor. Most studies on fermentative hydrogen production using mixed cultures have been conducted in conventional CSTR under mesophilic conditions. However, it is relatively difficult to achieve that in a CSTR because washout of the biomass usually occurs at a low hydraulic retention time (HRT). Attempts to enhance biomass retention by physical or biological immobilization of cells were shown to attain better hydrogen production performance than that of conventional CSTR, with hydrogen production rates ranging from 6–360 L/d/L [9–11]. Nevertheless, the matrices used for cell immobilization inevitably occupy significant space in the reactor, limiting cell density and possibly generating mass transfer barriers between
* Corresponding author. Tel.: þ86 10 62620903; fax: þ86 10 62559373. E-mail address: [email protected] (J. Shen). 0360-3199/$ – see front matter ª 2009 Professor T. Nejat Veziroglu. Published by Elsevier Ltd. All rights reserved. doi:10.1016/j.ijhydene.2009.04.018
international journal of hydrogen energy 35 (2010) 2622–2626
substrates and products. To avoid these problems, granular sludge was generated to enhance cell retention and biomass concentration simultaneously [12]. Biofilm-based systems were also used for anaerobic hydrogen production since they were more capable of maintaining higher biomass concentration and could operate at low HRT without biomass washout problems [13]. Some metal ions, such as calcium ion, were supplemented as coagulant aids to enhance the biomass accumulation [14]. However, the feasibilities of using the granular sludge or biofilm systems rely on the ability to control the timing and conditions of sludge granulation or biofilm formation as well as to maintain the stability and activity of granular sludge or biofilm. Furthermore, it is usually time-consuming to start up an up-flow anaerobic sludge blanket (UASB) or biofilm-based reactor, and poor startup of biological hydrogen production systems cause an ineffective hydrogen production rate and poor biomass growth at a high HRT, or cause a prolonged period of acclimation. An attractive and effective way may be that increasing the cell density in the reactor without immobilization matrices, as well as to ensure a relatively short start-up time and make the reactor easy to operate. In the consideration above, a conventional CSTR was utilized to study the maximization of hydrogen productivity using the well-mixed cultures used in our previous studies [6], owing to that CSTR is simple, easy to operate and low-cost without immobilization carrier. Furthermore, it usually takes shorter start-up time compared with UASB and biofilm systems. In order to improve the operation stability and maintain a high biomass density at relatively low HRT aiming to obtain a high hydrogen production rate as well as a high substrate utilizing rate, 100 mg/L calcium ions were added into the feed medium. The effect of calcium supplementation on the performance of CSTR was investigated by examining the hydrogen characteristics and metabolites distributions all over the operation course.
2.
Experimental section
2.1.
Inoculum, medium and analytical procedures
The inoculum of mixed cultures with mainly Clostridium pasteurianum bacteria, the synthetic medium with 11.1 g/L sucrose and the analytical procedures were the same as our previous work described in Ref [6].
2.2.
Operating conditions
The CSTR used in this study was 1 L in working volume with an internal diameter of 8 cm and a liquid height of 20 cm. The reactor was operated at 35 C and stirred by gas circulation. The HRT reduction was conducted in a stepwise manner through 78 days, respectively. At each HRT, the reactor was operated for about 2 weeks to allow steady-state condition development. Steady-state conditions were established when the variation in the product concentrations was constant (gas production rate, 5%; substrate degradation efficiency, 5%). After the constant parameter data were obtained, HRT was then shortened. Calcium with concentration of 100 mg/L was
2623
added in the feed medium at both 4 h and 2 h HRTs for comparison.
3.
Results and discussions
3.1. Reactor operation and hydrogen production characteristics Compared to UASB systems, which usually need more than 30 days to start up [12], it is time saving to start up a CSTR system. At an HRT of 8 h, the reactor achieved constant digestion gas production after 10 days of acclimation. The reactor needed less than 2 days to achieve constant digestion gas production after HRT reduction at each step except for the HRT reduction from 4 h to 2 h without calcium supplementation. The reactor was monitored by examining the effluent wastewater every three days for substrate degradation efficiency (Sd), hydrogen content (HC), hydrogen production rate (HPR), hydrogen yield (HY), volatile suspended solids (VSS), total suspended solids (TSS) and metabolites concentrations. As shown in Fig 1, the CSTR operated in a smooth state at HRTs of 8 h and 6 h. At both HRTs, the sucrose degradation efficiencies were over 98%. Gas product analyses indicated that the biogas only consisted of hydrogen and carbon dioxide, and the HCs were constantly 43% and 50% at HRT 8 h and HRT 6 h, respectively. The HPRs were 6.1 L/d/L and 10.0 L/d/L at HRT 8 h and HRT 6 h, respectively. From the VSS data we could see that the total amount of bacteria increased along with the decrease of the HRT, resulting in the increase of the hydrogen-producing bacteria and hydrogen production rate. When HRT decreased from 6 h to 4 h, sucrose degradation efficiency decreased to 89%, which was mainly due to the cell washout effect, because VSS value decreased from1054 at HRT 6 h to 889 mg/L at HRT 4 h caused by the increase of the substrate loading rate. Fortunately, the HC, HPR and HY all increased to the maxima of 55%, 14 L/d/L and 3.63 mol H2/mol sucrose, respectively. The reason for this phenomenon was probably that the content of hydrogen-producing bacteria in the mixed cultures increased along with the decrease of HRT, in spite of the decrease of total bacterial population. It was reported that hydrogen-producing bacteria reproduced much faster than other bacteria, so at relatively lower HRT, their content in the bacterial community usually increases with the decrease of HRT [15]. The reactor became quite unstable when HRT decreased to 2 h. All the parameters fluctuated largely during 15-day operation. Both the sucrose degradation efficiency and hydrogen content decreased to about 39%, which were the lowest during the whole operation course. HPR decreased to about 12.8 L/d/L, which was lower than that of 4 h HRT. Nevertheless, the hydrogen yield increased a little to 3.76 mol H2/mol sucrose, and was relatively constant despite of the fluctuations of Sd and HY. This was probably because that the ratio of the hydrogen production bacteria in the community was constant although the total cell density of the whole bacteria community changed with the environmental parameters [16]. In order to achieve a stable operation bioreactor at low HRT aiming to maximize the hydrogen productivity of hydrogen-producing bacteria, 100 mg/L calcium ions
2624
international journal of hydrogen energy 35 (2010) 2622–2626
Fig. 1 – Performance of CSTR: (a) hydrogen retention time (HRT), (b) substrate degradation efficiency (Sd) and hydrogen content (HC), (c) hydrogen production rate (HPR) and hydrogen yield (HY), (d) volatile suspended solids (VSS) and metabolites concentrations.
were added into the feed medium at HRT 2 h. When calcium was added into the medium at 2 h HRT, the substrate degradation efficiency increased to 73%, which was almost twice of that without calcium supplementation. HPR and HY increased to 24.5 L/d/L and 3.74 mol H2/mol sucrose, which were the highest values our research group has achieved and were relatively high compared with other hydrogen-producing bacteria in the CSTR mode [17,18]. Table 1 shows that VSS increased to 1682 mg/L, which was much higher than 747 mg/ L than that without calcium supplementation, indicating that cell retention in the reactor was the main reason for this improvement. Another reason was probably that calcium was required for catalytic activity, either as a participant in the reaction, or as a structural requirement in order to maintain the appropriate confirmation of the active site [12].
Interestingly, compared to UASB or biofilm-based systems with calcium supplementation [12,13], bacteria in our CSTR were always maintained in a suspended state and no granular sludge or biofilm formed during the whole operation, indicating an alternative way to improve performance of bioreactor by merely increasing the cell density, as well as avoiding the disadvantages of UASB or biofilm-based systems. As for comparison, with calcium supplementation at HRT 4 h, the sucrose degradation efficiency increased highly to 99%, and all the operation parameters almost remained constant during 15-day operation, indicating that the bioreactor became quite more stable mainly due to cell retention exhibiting the increase of VSS from 889 to 1092 mg/L. However, the HPR and HY decreased to about 11.2 L/d/L and 2.58 mol H2/mol sucrose, respectively. This might be that the
Table 1 – Substrate degradation efficiencies, biomass and hydrogen production characteristics. HRT (h) 8 6 4 2 4 2 a b c d e
Ca2þ (mg/L) 0 0 0 0 100 100
Sda (%)
HCb (%)
HPRc (L/d/L)
HYd (mol H2/mol sucrose)
99 1 99 1 89 4 38 9 99 73 5
43 50 55 39 3 42 1 47 2
6.1 0.1 10.0 0.2 14.0 1.3 12.8 2.5 11.2 24.5 0.5
2.80 0.01 3.48 0.03 3.63 0.12 3.76 0.22 2.58 3.74 0.32
Substrate degradation efficiency. Hydrogen content. Hydrogen production rate. Hydrogen yield. Ratio of volatile suspended solids and total suspended solids.
VSS (mg/L)
VSS/TSSe (%)
874 127 1054 80 889 152 747 167 1092 82 1682 470
87 93 86 90 73 72
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international journal of hydrogen energy 35 (2010) 2622–2626
Table 2 – pH values and metabolites distributions. HRT (h) 8 6 4 2 4 2 a b c d e f
Ca2þ (mg/L)
pH
0 0 0 0 100 100
5.70 0.05 5.40 0.19 5.44 0.12 6.47 0.11 5.16 0.27 5.35 0.26
EtOHa (mg/L) 760 80 610 10 510 120 1280 390 300 300 30
HAcb (mg/L)
HPrc (mg/L)
HBud (mg/L)
(A þ B)e (mg/L)
A/Bf
800 30 1340 20 1750 220 2370 230 570 140 2710 400
640 30 470 110 830 170 600 370 190 60 300 180
1060 40 1240 20 1730 230 1500 330 2260 10 1280 260
1860 2580 3480 3870 2830 3990
0.75 1.08 1.01 1.58 0.25 2.12
Ethanol. Acetate. Propionate. Butyrate. Sum of acetate and butyrate. Acetate/butyrate ratio.
bacterial community changed when calcium was added into the medium.
3.2.
Metabolites distributions
As for C. pasteurianum, which was dominant in a CSTR fermentor seeded with this kind of suspended mixed cultures, a theoretical maximum of 4 mol H2 per mol of hexose is obtained when acetic acid is the resulting product. When butyrate is the sole resulting product, a theoretical maximum of 2 mol H2 per mol of hexose is obtained [19]. In practice, high H2 yields are usually associated with a mixture of acetate and butyrate as fermentation products, and low H2 yields are associated with propionate and reduced resulting products (alcohols, lactic acid) [20]. Table 2 shows that the major products were butyrate, acetate, ethanol and propionate with average concentrations of 1060–2260, 570–2710, 300–1280 and 190–830 mg/L, respectively. As the HRT decreased from 8 h to 2 h without calcium supplementation, the sum of butyrate and acetate increased from 1860 to 3870 mg/L, which coincided with the increasing trend of HY. With the decrease of HRT from 8 h to 2 h, the acetate/butyrate ratio also showed an increasing trend except at HRT 4 h, which supported that higher concentration of acetate was benefit to hydrogen production. The decrease of acetate/butyrate ratio at HRT 4 h did not lower the HY, showing that the sum of acetate and butyrate determined higher HY under this condition. At HRT 2 h with calcium supplementation, the sum of acetate and butyrate and the acetate/butyrate ratio both increased to 3990 mg/L and 2.12 respectively, which were the highest during the whole operation course, which were consistent with the highest HY of 3.74 mol H2/mol sucrose. It proved again the conclusion that calcium supplementation at lower HRT improves the hydrogen productivity and alters the fermentation pathway to produce more hydrogen.
4.
Conclusions
The maximization of hydrogen productivity of suspended hydrogen-producing mixed cultures in a conventional CSTR was studied with calcium supplementation at low HRT. Operations of CSTR at HRT of 8–4 h without calcium
supplementation were stable with substrate degradation efficiencies more than 89%. The HPR and HY reached the peak at 14 L/d/L and 3.63 mol H2/mol sucrose at HRT 4 h, respectively. However, when the bioreactor was operated at a low HRT of 2 h, operation of bioreactor became quite unstable. Thus 100 mg/L calcium ions were added into the feed medium of HRT 2 h in order to maintain a high cell density in the fermentor. The bioreactor became more stable and the substrate degradation efficiency increased to 73%, which was twice that of bioreactor without calcium supplementation. HPR and HY were increased dramatically to 24.5 L/d/L and 3.74 mol H2/mol sucrose, respectively, which were the highest values we have ever reached. Retaining a high cell density and optimization of bacterial community in the bioreactor will be equally important to achieve higher hydrogen productivity and a better performance.
Acknowledgments The authors would like to thank the Ministry of Science & Technology, China for financial support under the National Hi-Tech R&D Program (863 Program, Grant No. 2006AA 05Z101).
references
[1] Lubitz W, Tumas W. Hydrogen: an overview. Chem Rev 2007; 107:3900–3. [2] Hallenbeck PC, Benemann JR. Biological hydrogen production: fundamentals and limiting processes. Int J Hydrogen Energy 2002;27:1185–93. [3] Kotay SM, Das D. Biohydrogen as a renewable energy resource-Prospects and potentials. Int J Hydrogen Energy 2008;33:258–63. [4] Lin CY, Lay CH. Carbon/nitrogen-ratio effect on fermentative hydrogen production by mixed microflora. Int J Hydrogen Energy 2004;29:41–5. [5] Van Ginkel SW, Oh SE, Logan BE. Biohydrogen gas production from food processing and domestic wastewaters. Int J Hydrogen Energy 2005;30:1535–42. [6] Yuan Z, Yang H, Zhi X, Shen J. Enhancement effect of l-cysteine on dark fermentative hydrogen production. Int J Hydrogen Energy 2008;33:6535–40.
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international journal of hydrogen energy 35 (2010) 2622–2626
[7] Ren N, Li J, Li B, Wang Y, Liu S. Biohydrogen production from molasses by anaerobic fermentation with a pilotscale bioreactor system. Int J Hydrogen Energy 2006;31: 2147–57. [8] Yu HQ, Mu Y. Biological hydrogen production in a UASB reactor with granules. II: reactor performance in 3-year operation. Biotechnol Bioeng 2006;94:988–95. [9] Wu SY, Hung CH, Lin CN, Chen HW, Lee AS, Chang JS. Fermentative hydrogen production and bacterial community structure in high-rate anaerobic bioreactors containing silicone-immobilized and self-flocculated sludge. Biotechnol Bioeng 2006;93:934–46. [10] Wu SY, Lin CN, Chang JS. Hydrogen production with immobilized sewage sludge in three-phase fluidized-bed bioreactors. Biotechnol Prog 2003;19:828–32. [11] Lee KS, Lo YC, Lin PJ, Chang JS. Improving biohydrogen production in a carrier-induced granular sludge bed by altering physical configuration and agitation pattern of the bioreactor. Int J Hydrogen Energy 2006;31:1648–57. [12] Yu H, Zhu Z, Hu W, Zhang H. Hydrogen production from rice winery wastewater in an upflow anaerobic reactor by using mixed anaerobic cultures. Int J Hydrogen Energy 2002;27: 1359–65. [13] Ahn Y, Park EJ, Oh YK, Park S, Webster G, Weightman AJ. Biofilm microbial community of a thermophilic trickling
[14]
[15]
[16]
[17]
[18]
[19]
[20]
biofilter used for continuous biohydrogen production. FEMS Microbiol Lett 2005;249:31–8. Kim IS, Jang N. The effect of calcium on the membrane biofouling in the membrane bioreactor (MBR). Water Res 2006;40:2756–64. Sahlstrom L. A review of survival of pathogenic bacteria in organic waste used in biogas plants. Bioresour Technol 2003; 87:161–6. Chang FY, Lin CY. Biohydrogen production using an up-flow anaerobic sludge blanket reactor. Int J Hydrogen Energy 2004; 29:33–9. Gavala HN, Skiadas IV, Ahring BK. Biological hydrogen production in suspended and attached growth anaerobic reactor systems. Int J Hydrogen Energy 2006;31:1164–75. Kim SH, Han SK, Shin HS. Performance comparison of a continuous-flow stirred-tank reactor and an anaerobic sequencing batch reactor for fermentative hydrogen production depending substrate concentration. Water Sci Technol 2005;52:23–9. Kleerebezem R, Stams AJ. Kinetics of syntrophic cultures: a theoretical treatise on butyrate fermentation. Biotechnol Bioeng 2000;67:529–43. Vignais PM, Magnin JP, Willison JC. Increasing biohydrogen production by metabolic engineering. Int J Hydrogen Energy 2006;31:1478–83.