Exploring biohydrogen-producing performance in three-phase fluidized bed bioreactors using different types of immobilized cells Shu-Yii Wu1*, Chi-Neng Lin1, Yuan-Chang Shen1, Chiu-Yue Lin2 and Jo-Shu Chang3 1
Dept. of Chemical Engineering, Feng Chia University, Taichung, Taiwan
2
Dept. of Environmental Engineering & Science, Feng Chia University, Taichung, Taiwan 3
Dept. of Chemical Engineering, National Cheng Kung University, Tainan, Taiwan
*
Research Center for Energy and Resources, Feng Chia University, Taichung, Taiwan *
E-mail:
[email protected]
ABSTRACT In this study, the spherical activated carbon (AC) and silicone gel (SC) were used as the primary matrices to immobilize H2-producing activated sludge. The experiments were carried out in two different types of three-phase fluidized beds; namely, conventional fluidized bed reactor (FBR) and draft tube fluidized bed reactor (DTFBR). The solid volume of AC and SC immobilized cells was 10 vol.% for both FBR and DTFBR. Sucrose (at 20000 mg COD/l) was used as the carbon substrate for H2 production. The H2-producing performance was examined at different hydraulic retention times (HRT = 8, 6, 4, 2, 1, and 0.5 h). The results show that the best volumetric H2 production rate was 1.23 ± 0.08 l/h/l (HRT = 2 h) and 2.33 ± 0.22 l/h/l (HRT = 0.5 h) for fluidized beds containing AC and SC immobilized cells, respectively. The highest H2 yield was 3.37 mol H2/mol sucrose (HRT = 6 h) and 4.07 mol H2/mol sucrose (HRT = 4 h) for fluidized beds with AC and SC immobilized cells, respectively. The H2 content in the biogas was stably maintained at 35% or higher for all the reactors, while the primary soluble metabolites in the cultures were acetic acid and butyric acid.
Keywords: Biohydrogen, Activated carbon (AC), Silicone gel (SC), Anaerobic sludge, Fluidized bed reactor (FBR), Draft tube fluidized bed reactor (DTFBR) 1. INTRODUCTION Immobilized-cell systems have been successfully applied in various bioreactors for biohydrogen production [1-3].
Using immobilized-cell systems could increases the
concentration and retention of biomass, allowing stable operation at high dilution rates (HRT < 4h). These features led to marked improvement in the hydrogen production rate. Three types of immobilized-cell systems have been applied in biohydrogen production, including surface attachment [1], self-flocculation [2], and gel entrapment [3] approaches, among which the surface attachment approach was most frequently used for dark H2 fermentation. Moreover, activated carbon (AC) was the most common matrix for cell growth and biofilm attachment for H2 production [4], while fibrous support matrix [5] and cellulosic matrix [6] were also used.
These matrices are readily available and easy to handle, but the biofilms were usually not firmly bound on the surface of carrier supports and could fall off due to shear force or hydraulic pressure. In contrast, cell entrapment causes mass transfer limitation, but allows higher biomass content of H2-producing bacteria in the reactors and creates a local anaerobic environment, which is well suited to oxygen-sensitive fermentative H2 production. Therefore, this work attempted to use both spherical activated carbon (AC) and silicone-immobilized cells (SC) to retain higher biomass for high-rate hydrogen production in fluidized bed reactor (FBR) and draft tube fluidized bed reactor (DTFBR), converting sucrose substrate to H2 during continuous operation. The FBR and DTFBR were operated at different HRT (8, 6, 4, 2, 1 and 0.5 h) to investigate the effect of HRT on the H2-producing ability. 2. MATERIALS AND METHODS
2.1. H2-producing sludge The anaerobic sludge obtained from a municipal sewage treatment plant in Taichung, Taiwan was used as the seed sludge. The typical pH, volatile suspended solid (VSS) and total solid (TS) concentration of the sludge were 6.81, 33.3 and 65.1 g/l, respectively. The hydrogen productivity of the seed sludge was enhanced by thermal treatment at 100 oC for 1 h to inhibit the methanogenic activity. The thermally treated sludge was further acclimated in a continuous culture operated at HRT = 8 h with an influent sucrose concentration of 20000 mg COD/l [7]. The effluent sludge was collected for cell immobilization. 2.2. Medium compositions The medium used for H2 fermentation consisted of sucrose as the sole carbon substrate (initial sucrose concentration (CSo) = 20000 mg COD/l) and sufficient inorganic supplements, including (mg/l) NH4HCO3, 5240; NaHCO3, 6720; K2HPO4, 125; MgCl2·6H2O, 100; MnSO4·6H2O, 15; FeSO4·7H2O, 25; CuSO4·5H2O, 5; CaCl2, 100 and CoCl2·5H2O, 0.125. Sucrose is a very common and effective substrate for fermentative H2 production, leading to great hydrogen productivity [1-4, 7, 8]. 2.3 Immobilization of H2-producing sludge 2.3.1 Cell immobilization on spherical activated carbon Hydrogen-producing seed sludge (3-5 g-VSS/l) was cultivated in a 2.5 L fluidized bed reactor (FBR) containing spherical activated carbon (size: 2-3 mm) with a solid volume of 10 vol%. The reactor was initially operated on batch mode for 48 h to allow biofilm formation on the surface and macropores of AC.
2.3.2 Cell Immobilization with silicone gel Hydrogen-producing sludge was mixed with activated carbon powder (Union Chemical Works Ltd., Hsinchu, Taiwan) at a volume (ml) to weight (g) ratio of 10:1. The sludge/activated carbon slurry was then mixed with silicone gel (SC) (weight to weight ratio = 1:10). The final mixture was then extruded to produce SC-immobilized sludge, which was disc shaped with a 3
height, diameter, and density of 0.3 cm, 0.4 cm, and 1.31 g/cm , respectively. The detailed procedures of preparing the immobilized sludge were described in our previous study [3]. 2.4. Set-up and operation of FBR/DTFBR for H2 production Schematic descriptions of the three-phase fluidized beds containing AC- and SC-immobilized cells are shown in Fig. 1 and Fig. 2. The experiments were carried out in two different types of three-phase fluidized beds; namely, conventional fluidized bed reactor (FBR) and draft tube fluidized bed reactor (DTFBR). The main body of the two reactors was a cylindrical column with 8 cm in diameter and 24 cm in height (working volume = 2.5 L). The major difference between the two bioreactors was that DTFBR contained a drift tube inside the column, avoiding the overflow of floating immobilized-cell particles from the upper part of the column. The draft tube can also improve the separation of the biomass and the biogases in the exit stream. For FBR system, the recycle rate was controlled at 5.5 L/min (bed expansion= 10%) maintaining well fluidization of the particles in the reactor (Fig.1). While the DTFBR was controlled at 3 L/min, it also kept well fluidization in the reactor (Fig.2). Both bioreactors were initially operated on batch mode for 48 h to activate the H2-producing sludge before they were switched to a continuous mode at a designated hydraulic retention time (HRT = 8, 6, 4, 2, 1 and 0.5h). The effluent of the reactor went to a gas-liquid separator, where the gaseous and soluble products were collected separately (Fig.1 and Fig.2). After reaching steady-state operation (based on a constant volumetric H2 production rate with a variation of within 5-10% for 3-5 days), the HRT was decreased progressively from 8 to 0.5 h. The compositions of gas products (H2 and CO2) and soluble metabolites (volatile fatty acids, alcohols, etc.) produced during H2 fermentation was monitored as a function of time. The pH and biomass concentration (represented by volatile suspended solid; VSS) in the effluent were also recorded (Fig.1 and Fig.2). The reactor was operated at a temperature of 40oC, and a pH of within 6-7. A gas meter (Type TG1; Ritter Inc., Germany) was used to measure the amount of gas products generated and the gas volumes were calibrated to 25 oC and 760 mmHg. 2.5. Analytical methods Hydrogen gas was determined with a gas chromatography (GC-14A, Shimadzu, Tokyo, Japan) equipped with a thermal conductivity detector (TCD). The carrier gas was argon and the column was packed with Porapak Q (80/100 mesh, Waters Corp., USA). The VFA and
ethanol were also detected by gas chromatography (Shimadzu GC-14A) using a flame ionization detector (FID).
The carrier gas was N2 and the packing material was FON
(Shimadzu, Japan). Volatile suspended solid (VSS; representing the biomass concentration) was measured according to the procedures described in Standard Methods [9]. The sugar concentration in the effluent was also determined according to Standard Methods.
Fig. 1 Schematic description of the FBR for hydrogen fermentation
Fig. 2 Schematic description of the DTFBR for hydrogen fermentation
3. RESULTS AND DISCUSSIONS 3.1 Effect of HRT on H2 production in FBR/AC-immobilized cell (AC-FBR) system After starting up at a HRT of 8 h, the H2 production rate and H2 content in biogas were 0.52 l/h/l and 41.4%, respectively (Table 1). When the operations reached steady state, the HRT were adjusted gradually from 8 h to 6, 4 and 2 h. The H2 production rate increased with decreasing HRT. In particular, when the HRT was shifted down from 8 to 6 h, the organic loading rate increased 30%, while the H2 production rate increased 50% (from 0.52 to 0.78 l/h/l). Apparently, the H2 production was dependent on organic loading rate. However, when HRT was decreased to I h, the AC-FBR system encountered cell wash-out, the H2 production rate and H2 content decreased dramatically, resulting in system disruption. The results show that when the organic loading rate exceeded certain level (i.e. HRT = 1 h), the AC-FBR system could not be stably operated even with the AC-immobilized cells. This instability was in part due to severe overflow of the AC particles while operating at HRT = 1 h. The highest H2 production rate (1.23 l/h/l) occurred when AC-FBR was operated at 2 h HRT, while the highest H2 yield of 3.37 mol H2/mol sucrose was obtained at a HRT of 6 h (Table 1). The substrate conversion and H2 content were essentially over 90% and 40%, respectively, except operation at HRT = 1 h. Although the H2 production tended to increase as HRT decreased, the H2 yield and substrate conversion decreased with a decrease of HRT from 6 to 2 h.
This decrease was accompanied by a decrease in suspended biomass
concentration in the system. This seems to suggest that both suspended and immobilized cells contributed to H2 production. When HRT decreased, the suspended cell concentration also decreased, leading to a decrease in substrate conversion and H2 yield. As a result, the bacterial populations on the biofilms (Fig. 3) became the major H2 producers in the AC-FBR system. While operation at 1-h HRT, both suspended and AC-immobilized cells were washed out, resulting in termination of H2 production. For all experiments in AC-FBR system, the biogas contained only H2 and CO2, while methane was not detected. 3.2 Effect of HRT on H2 production in DTFBR/SC-immobilized cell (SC-DTFBR) system Our previous study showed [10] that the SC immobilized cells were very effective in hydrogen production in a batch or CSTR system. Table 1 showed the H2-producing performance when the SC-immobilized cells were applied in DTFBR system (SC-DTFBR system). The results showed that at HRT = 8 h, the H2 production rate and H2 content were 0.61 l/h/l and 43.6%, respectively. Increasing HRT from 8 to 4 h gave rise to an 85% increase in H2 production rate and the highest H2 yield of 4.07 mol H2/mol sucrose were obtained at 4-h HRT. Further decrease in HRT (from 4 to 0.5 h) elevated the H2 production rate from 1.13 to 2.33 l/h/l, whereas both H2 content and H2 yield decreased sharply (Table 1). Meanwhile, the substrate conversion also decreased to 75% when HRT was decreased to 0.5 h. It is worth noting that the SC-DTFBR system was able to operate stably without wash-out, indicating that
the design of drift tube in DTFBR system seemed to be effective in avoiding the loss of SC-immobilized cells and thus enhanced the operational stability. Nevertheless, like in AC-FBR system, suspended cells were removed significantly when HRT was shorter than 4 h. This suggests that the H2 production in the SC-DTFBR system was essentially contributed by the immobilized cells at HRT ≤ 4 h. The H2 production rate was maintained or was even higher for a HRT shift-down of 4 to 0.5 h, whereas the H2 yield and substrate conversion dropped significantly with a lower cell concentration in the suspended phase. The biogas produced from the SC-DTFBR process contained only H2 and CO2, while methane was not detected during the course of experiments.
Table 1
Performance of dark H2 fermentation under different operation conditions in FBR (with AC immobilized cells) and DTFBR (with SC immobilized cells)
Bioreactor systems
AC-FBR
SC-DTFBR
HRT (h)
pH
CH2 (%)
HPR (l/h/l)
Biomass* (g-VSS/l)
Yield (mol H2/mol sucrose)
X (%)
8 6 4 2 1 8 6 4 2 1 0.5
6.4 6.4 6.7 6.9 WO 6.7 6.5 6.6 6.9 7.0 7.0
41.4 42.6 41.0 39.7 WO 43.6 43.2 44.2 35.0 37.1 35.8
0.52 ± 0.02 0.78 ± 0.04 0.93 ± 0.05 1.23 ± 0.08 WO 0.61 ± 0.02 0.80 ± 0.03 1.13 ± 0.12 0.66 ± 0.06 1.40 ± 0.12 2.33 ± 0.22
2.57 2.85 2.58 2.21 WO 2.33 2.21 2.02 1.36 1.81 0.45
2.95 3.37 3.01 2.10 WO 3.90 3.90 4.07 1.52 1.31 1.22
98.7 97.9 91.3 90.7 WO 99.1 98.1 88.1 66.0 81.2 75.4
HTR: hydraulic retention time; CH2: H2 content in biogas; HPR: H2 production rate; X: substrate conversion; WO: wash out; * biomass stands for suspended cells only.
AC-FBR
SC-DTFBR
Fig. 3 Photos taken for the culture of AC-FBR (HRT = 2 h) and SC-DTFBR (HRT = 0.5 h).
3.3 Comparison of H2-producing performance between AC-FBR and SC-DTFBR systems In this study, two H2-producing bioreactor systems (namely, AC-FBR and SC-DTFBR) were used to improve cell retention at high hydraulic pressure and to enhance the H2 producing performance. Our recent work showed that AC and SC immobilized cells can induce formation of granular sludge, leading to drastic enhancement on H2 producing performance, especially for HRT less than 4 h [2-5, 10]. However, due to using a high recycle rate (5.5 l/min for AC-FBR: and 3 l/min for SC-DTFBR), the granular sludge did not form in both reactors. Therefore, it is observed that the H2 producing activity was dominated by bacterial populations in the immobilized cells, while the suspended cells were significantly removed by hydraulic dilution for HRT < 4 h, indicated by the sharp decrease in biomass concentration in suspended phase (Table 1). However, the results show that both AC-FBR and SC-DTFBR systems can be stably operated at a low HRT of 2 h, while SC-DTFBR exhibited better biomass retention (Fig. 3) as it can be operated at an extremely low HRT of 0.5 h, in contrast to 2-h HRT for AC-FBR system. This may be attributed to the design of drift tubes in DTFBR system, preventing overflow of the immobilized cells. As the SC-immobilized cells can be retained in the reactor, the system could survive operation at a HRT of 0.5 h with a high H2 production rate of 2.33 l/h/l. It is also evidential from Table 1 and Fig. 4 that the SC-DTFBR system displayed better H2 production rate than the AC-FBR system at all HRT used. The H2 yield obtained from SC-DTFBR was also higher than AC-FBR for HRT = 4-8 h. This seems to indicate that cell immobilization via gel entrapment gave better performance than that via surface attachment for the use in fluidized bed reactors.
Furthermore,
considering all the performance indexes (rate, yield and conversion), it would be preferable to operate both systems at a HRT of about 4 h.
H2 Production Rate (l/h/l)
2.5
2.0
1.5
1.0
0.5
SC-DTFBR
AC-FBR
8
Bioreactor System s Fig. 4 Effects of bioreactor systems and HRT on H2 production rate
6
4
2
1
0.5
T HR
(h
)
3.4 Soluble metabolites and carbon substrate conversion In both AC-FBR and SC-DTFBR systems, the carbon substrate (sucrose) conversion was above 88% for HRT > 4 h, indicating effective consumption of the substrate. When HRT was lower than 4 h, the AC-FBR system still displayed high substrate conversion (> 90%), whereas the conversion in the SC-DTFBR system decreased sharply to 66-81%. This decrease was related to the decrease in biomass content in the suspended phase, as the biomass content in SC-DTFBR dropped to 0.45-1.36 g-VSS/l while operating at a HRT of 0.5-2 h.
This indicates
that the suspended cells played an important role in substrate utilization. It is also noted that irregardless of bioreactor systems, both substrate conversion and H2 yield decreased when suspended biomass content decreased. The major soluble metabolites in both bioreactor systems were acetic acid (HAC) and butyric acid (HBu), accounting for 20-27% and 43-56% of total soluble microbial products (SMP). From Table 2, at the same HRT, although AC-FBR had higher total volatile fatty acids (TVFA) and TVFA/SMP ratio than SC-DTFBR, the latter system attained higher HBu/HAC ratio (2.48-2.85) than the former system (2.12-2.36), except HRT = 2 h in SC-DTFBR. These results support that the SC-DTFBR system gave better hydrogen production rate than AC-FBR. In addition, the soluble metabolites of propionic acid (HPr) and ethanol (EtOH) were 8-23% and 4-7% of SMP respectively in the AC-FBR system, while for SC-DTFBR, the HPr/SMP and EtOH/SMP ratios were 10-15% and 9-19%, respectively. As both HPr and EtOH products were disadvantageous to hydrogen production [11-12], the lower HPr and EtOH production suggests that the cultures in both bioreactor systems carried out metabolic pathways in favor of H2 production, as the predominant metabolites were HBu and HAc (combined as 70-80% of SMP).
Table 2 Formation of soluble metabolites during H2 fermentation in two different bioreactor systems EtOH/ HAc/ HPr/ HBu/ HBu/ TVFA/ TVFA SMP SMP SMP SMP SMP HAc SMP (mg COD/l) (mg COD/l) (%) (%) (%) (%) (%) 8 8 22 18 52 2.36 92 7103 7684 6 7 21 23 49 2.33 93 7873 8510 AC-FBR 4 7 22 20 51 2.32 93 7523 8089 2 4 25 18 53 2.12 96 6803 7086 8 9 22 13 56 2.55 91 7251 7968 6 10 23 10 57 2.48 90 7077 7863 SC-DTFB 4 11 20 12 57 2.85 89 5894 6934 R 2 19 27 11 43 1.59 81 4998 6170 1 17 24 15 44 1.83 83 6509 7842 0.5 12 22 13 53 2.41 88 6255 7108 HAc: acetic acid; HPr: propionic acid; HBu: butyric acid; EtOH: ethanol; TVFA = HAc+ HPr+ HBu;
Bioreactor systems
HRT (h)
SMP = TVFA + EtOH
4. CONCLUSIONS This study compared the performance of two H2-producing fluidized bed bioreactors containing gel-entrapped or surface-attached immobilized cells as the primary H2 producers. Although suspended biomass concentration considerably decreased at HRT < 4 h, the presence of immobilized cells still provides H2 production activity, leading to stable operations at a HRT of 2 and 0.5 h for AC-FBR and SC-DTFBR, respectively. The SC-DTFBR system with SC-entrapped cells and the design of drift tube appeared to exhibit better performance in terms of H2 production rate, H2 yield and operational stability at most of HRT used. The SC-DTFBR system attained the best hydrogen yield and H2 production rate of 4.07 mol H2/mol sucrose (HRT = 4h) and 2.33 l/h/l (HRT = 0.5 h), respectively. The H2 content in the biogas was stably maintained at 35 % or higher in all experiments. Irregardless of the type of bioreactor systems, the dominant soluble metabolites in the cultures were acetic acid and butyric acid. The proposed three-phase fluidized bed bioreactors showed promising H2 producing performance and may have a potential to apply in practical biohydrogen production processes.
ACKNOWLEDGEMENT The authors gratefully acknowledge the financial support by National Science Council of Taiwan, R.O.C. (grant no. NSC 93-2211-E-035-014 & NSC-93-2214-E-035-002), by Bureau of Energy, MOEA, Taiwan ROC (grant no. 94-D0137-2 & 95-D0137-2) and by Feng Chia University (grant no. FCU-89-J040).
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