Effect of temperature and temperature shock on the stability of continuous cellulosic-hydrogen fermentation

Bioresource Technology 142 (2013) 304–311

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Effect of temperature and temperature shock on the stability of continuous cellulosic-hydrogen fermentation Samir I Gadow a,b, Hongyu Jiang a, Ryoya Watanabe c, Yu-You Li c,d,⇑ a

Department of Environmental Science, Graduate School of Environmental Studies, Tohoku University, Aoba-ku, Sendai 9808579, Japan Department of Agricultural Microbiology, Agriculture and Biology Research Division, National Research Center, Dokki, Cairo, 12311, Egypt c Department of Civil and Environmental Engineering, Graduate School of Engineering, Tohoku University, Aoba-ku, Sendai 9808579, Japan d Key Laboratory of Northwest Water Resource, Environment and Ecology, Ministry of Education, Xi’an University of Architecture and Technology, Xi’an 710055, China b

h i g h l i g h t s  The 55 ± 1 and 80 ± 1 °C presented long-term stable cellulosic-hydrogen fermentation.  The first study on the effect of temperature shock on bioH2 production of cellulose.  The reactor under 55 or 80 °C appeared to be more resilient to the shock than 37 °C.  The recovery period after the 2nd shock was shorter than after the 1st shock.

a r t i c l e

i n f o

Article history: Received 15 March 2013 Received in revised form 24 April 2013 Accepted 25 April 2013 Available online 13 May 2013 Keywords: Bio-hydrogen Cellulose Dark fermentation Temperature shock Continuous mode

a b s t r a c t Three continuous stirred tank reactors (CSTR) were operated under mesophilic (37 ± 1 °C), thermophilic (55 ± 1 °C) and hyper-thermophilic (80 ± 1 °C) temperatures for 164 days to investigate the effect of temperature and temperature shock on the cellulosic-dark hydrogen fermentation by mixed microflora. During steady state condition, the sudden decreases in the fermentation temperature occurred twice in each condition for 24 h. The results show that the 55 ± 1 and 80 ± 1 °C presented stable hydrogen yields of 12.28 and 9.72 mmol/g cellulose, respectively. However, the 37 ± 1 °C presented low hydrogen yield of 3.56 mmol/g cellulose and methane yield of 5.4 mmol/g cellulose. The reactor performance under 55 ± 1 or 80 ± 1 °C appeared to be more resilient to the sudden decreases in the fermentation temperature than 37 ± 1 °C. The experimental analysis results indicated that the changing in soluble by-products could explain the effect of temperature and temperature shock, and the thermophilic temperature is expected having a better economic performance for cellulosic-hydrogen fermentation. Ó 2013 Elsevier Ltd. All rights reserved.

1. Introduction Compared to other current fuel to energy conversion technologies, the higher efficiency of the conversion of hydrogen to electrical energy due to its high calorific value makes hydrogen a potential substitute for fossil fuels. Besides being energy efficient, it is also carbon-free, non-polluting, and recyclable. Over 95% of world’s hydrogen demand is now being derived from fossil fuels (Ewan and Allen, 2005). This presents a major problem since the same amount of CO2 as is formed in the combustion of fossil fuels is released (Zhao and Yu, 2008). Biological processes have the potential to generate hydrogen from renewable and/or recyclable feed-stocks. Over the past decade, the feasibility of bio-hydrogen ⇑ Corresponding author. Address: Dept. of Civil and Environmental Engineering, Graduate School of Engineering, Tohoku University, Aoba-ku, Sendai 980-8579, Japan. Tel.: +81 22 7957464; fax: +81 22 7957465. E-mail address: [email protected] (Y.-Y. Li). 0960-8524/$ - see front matter Ó 2013 Elsevier Ltd. All rights reserved. http://dx.doi.org/10.1016/j.biortech.2013.04.102

production from organic waste streams has been demonstrated (Lee et al., 2010). However, few studies have been dedicated to describing biological hydrogen production using cellulose. Because cellulose is particularly difficult to hydrolyze, its uses have been considered extremely limited (Lynd et al., 2002). Mainly, dark hydrogen fermentation commonly conducted under mesophilic temperature; however, under thermophilic temperature or higher, there are several advantages such as increased pathogenic destruction of residues, reduction methanogenic bacteria (Sahlstrom, 2003) and better thermodynamic conditions (Van Niel et al., 2002). Anaerobic processes are frequently believed to be sensitive to sudden changes in operational parameters (Speece, 1983). Therefore, the unexpected changes in the operating parameters such as sudden temperature (Temper et al., 1983), substrate concentrations variations (Angenent et al., 2002) and to change in substrate (Berg and Lentz, 1981) results unbalanced in the fermentation process. The effect of the temperature shock depends on the temperature itself, the exposure time and the bacterial

S.I Gadow et al. / Bioresource Technology 142 (2013) 304–311

composition of the sludge (Obaya et al., 1994). No detailed of the effects of temperature shock on biological hydrogen production has yet to appear in the literature. However, the effects of temperature shocks on the mesophilic and thermophilic anaerobic digestion of methane have been recently studied (Van Lier et al., 1990 and Ahring et al., 2001). In anaerobic reactors operated under the mesophilic conditions, temperature shock tends to result in unstable performance and in some extreme cases; it results in complete failure (Van Lier et al., 1990). Since the thermophilic range is considered more susceptible to environmental and operational conditions than the mesophilic condition (Ahring et al., 2001); some studies should be focused on the thermophilic or extreme thermophilic ranges. In the anaerobic process, the changes in the soluble by-products such as acetic acid or total volatile fatty acid concentrations have been considered an important indicator of instability under stressed conditions (Graef and Andrews, 1974). In industry, it is important for systems to be able to cope with such unexpected interruptions as maintenance and power failure. The primary impact of temperature on microbial activity and community composition in a wastewater treatment reactor has been well established (Gao et al., 2011). However, there are some causes of temperature variations such as operational or environmental conditions so that the system can be adjusted to accommodate to the new conditions, whereas sudden transient changes can lead to a marked deterioration in the reactor’s performance (Gao et al., 2011). Therefore, the purposes of this study were to examine the effect of fermentation temperature and temperature shock on the continuous cellulosichydrogen fermentation without prior pretreatment and to evaluate the long-term stability of fermentation performance under steady state condition.

2. Methods 2.1. Seed microflora and substrate The anaerobic mixed microflora was obtained from a sewage sludge digester at the Sendai municipal sewage treatment plant in Japan without pretreatment. The medium for dark H2 fermentation was constituted of the following [g/L]: Cellulose 10 (Cellulose powder E, Toyo Roshi Kaisha, Ltd. Japan), NH4HCO3 2, NaHCO3 3, K2HPO4 0.125, MgCl26H2O 0.1, MnSO46H2O 0.015, FeSO47H2O 0.025, CuSO45H2O 0.005 and CoCl25H2O 1.25  104.

305

2.2. Experimental procedure The experimental apparatus was composed of two parts, including feedstock tank and H2 producing acidogenic reactor. A substrate maintained at 4 °C in feedstock tank was fed into CSTR by a peristaltic pump with a time-control system (see Fig. 1). The CSTR were operated under different temperatures, 37 ± 1 °C (mesophilic), 55 ± 1 °C (thermophilic) and 80 ± 1 °C (hyperthermophilic) with an effective volume of 6 L and operated using a hydraulic retention time [HRT] of 10 days.

2.3. Temperature shock The operational temperature was controlled by circulating water through the water jacket. Temperature shock occurred twice during the steady-state conditions at each condition for 24 h. The temperature was varied by adjusting the temperature of the surrounding water baths. The reactors were subjected to periodic temperature shock: 11, 23 and 44 °C shock (from 37 to 26 °C, 55 to 32 °C and 80 to 36 °C) at mesophilic, thermophilic and hyperthermophilic temperatures, respectively every time.

2.4. Analytical methods The percentage of H2 in the biogas was determined by a gas chromatograph (Shimadzu 8A) equipped with a thermal conductivity detector (TCD) and a stainless steel column packed with a molecular sieve 5A (60/80 3 mmØ). The temperatures of the detector and the column were maintained at 100 and 60 °C, respectively. For the determination of the carbon dioxide, nitrogen and methane content, the same model of gas chromatograph (Shimadzu 8A) was used, and it was equipped with a TCD and a stainless steel column packed with Porapak T was used. The temperatures of the detector and the column, here, were maintained at 100 and 70 °C, respectively. The organic acids and ethanol were analyzed by a gas chromatograph (Shimadzu GC-1700) equipped with a flame ionization detector (FID) and a 30 m column (J&W DB-WAX). The volatile suspended solid (VSS), volatile solid (VS), protein and chemical oxygen demand (COD) were measured according to the procedures described in the Standard Methods (APHA, 2005).

Fig. 1. Schematic diagram of experimental apparatus. 1 – Feedstock tank, 2 – Feed inlet, 3 – Mixer, 4 – Recirculation cooler, 5 – Sampling pump, 6 – Hydrogen reactor, 7 – Hot water recirculation, 8 – Thermometer, 9 – Gas-water separation chamber, 10 – Wet gas meter, 11 – Sample port, 12 – Effluent pump, 13 – Digestion sludge tank.

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S.I Gadow et al. / Bioresource Technology 142 (2013) 304–311 2 nd Shock

1 s t Shock

Gas content %

Gas production l/l/day

0.6 0.5

Steady state Recovery condition duration

Steady state condition

Steady state condition

Recovery duration

0.4 0.3

(a)

0.2 40

CH4 H2

30 20

(b)

10

Total VFA g/L

pH

0

7 6 5 4 3 4 3 2 1

(c)

(d)

VSS g/L

8 6 4

(e)

2 90

100

110

120

130

140

150

160

Operation time (days) Fig. 2. Evolutions of biogas production rates (a), gas concentrations (b), pH (c), total Volatile fatty acid (VFA) g/L (d), and VSS g/L (e) in the mesophilic reactor.

3. Results and discussion 3.1. Time course profiles of three bioreactor performance under steady state condition 3.1.1. Mesophilic temperature Through 160 days of operation, the total biogas production, gases concentrations, pH, total volatile fatty acid (VFA) and volatile suspended solid (VSS) were analyzed to investigate the influence of the temperature and temperature shock on bioH2 of cellulose at the mesophilic condition (see Fig. 2). Under the steady state condition, as much as to 0.43 L/L/day biogas, with a gas composition of 18.5%, 28.2% and 53.3% hydrogen, methane and carbon dioxide gases, respectively was produced as shown in Fig. 2(a and b). The results confirmed that the untreated seed sludge by thermal pretreatment was responsible for generating methane gas in the gas content as reported in previous studies using cellulose in the batch (lay, 2001 and Kumar and Lin, 2013) and continuous mode (Gadow et al., 2012). The pH in the reactor was maintained of 5.6 ± 0.16 (Fig. 2(d)) with the use sodium bicarbonate, which is known for its strong buffering of growth medium (Gadow et al., 2012). At the first shock, biogas production decreased to 0.32 L/L/day, with a gas composition of 16%, 27% and 57% hydrogen, methane and carbon dioxide gases, respectively. No significant change in the pH was observed. Once the temperature was recovered, the system successfully returned to the steady state condition with biogas rate of 0.41 L/L/day, and gas composition of 16.75%, 27.3% and 56%

hydrogen, methane and carbon dioxide gases, respectively. On the other hand the detrimental effect of the second shock on the system performance in terms of gas composition was observed. The biogas production decreased to 0.32 L/L/day, with 10%, 25% and 65% hydrogen, methane and carbon dioxide, respectively. The system again reached a steady state condition, with biogas production of 0.36 L/L/day and composition of 6.1%, 30.2% and 63.7% hydrogen, methane and carbon dioxide gases, respectively. The results are clear evidence that the temperature shocks result in the unstable performance of hydrogen production under mesophilic temperature; however, there is no considerable change in term of methane production. 3.1.2. Thermophilic temperature The reactor was operated at 55 ± 1 °C for over five months to ensure a steady state condition for the total biogas production, H2 concentrations, pH, VFA and VSS (see Fig. 3). Under the steady state condition, the system produced 0.5 L/L/day biogas, with 55% and 45% hydrogen and carbon dioxide gases, respectively as shown in Fig. 3(a and b) and no methane was detected. Under such condition, the pH was maintained at 5.8 ± 0.1 throughout the steady state with the use sodium bicarbonate (see Fig. 3(d)). The results show that the 55 °C and pH inhibited methanogenic bacteria during cellulose fermentation by mixed culture compared with mesophilic temperature (Luo et al., 2011). These results suggest that the main group of sporeforming anaerobes present in the cellulose-enriched culture started from sewage sludge appeared

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Steady state condition

0.2

Steady state condition

(a)

0.0 60

Gas production L/L/day

Steady state Recovery condition duration

0.4

0.2

40

Steady state condition

(a)

40

(b)

20

(b) pH

0 7

6 5 4 3 2

(d)

1

VSS g/L

0 8 6 4 2 0

(e) 90

100

110

120

130

140

150

160

Operation time (days)

6

(c)

5

(c) Total VFA g/L

pH

Steady state condition

0.4

0 7

Total VFA g/L

Steady state Recovery condition duration

0.0 60

20

VSS g/L

2 nd Shock

Recovery duration

0.6

H2%

H 2%

Gas production L/L/day

Recovery duration

0.6

1 s t Shock

2 nd Shock

1 s t Shock

4 3 2 1 0 8 6 4 2 0

(d)

(e) 90

100

110

120

130

140

150

160

Operation time (days) Fig. 4. Evolutions of biogas production rates (a), H2 concentrations (b), pH (c), Total Volatile fatty acid (VFA) g/L (d) and VSS g/L (e) in the hyper-thermophilic reactor.

Fig. 3. Evolutions of biogas production rates (a), H2 concentrations (b), pH (c), Total Volatile fatty acid (VFA) g/L (d) and VSS g/L (e) in the thermophilic reactor.

to be cellulosic hydrogen bacteria. The system was exposed to temperature shock (from 55 to 32 °C), resulting in a 0.5–0.2 L/L/ day decrease in biogas production with shift 55–30% in the hydrogen gas. It should be noted that the pH increased to 6.4 during the recovery period, perhaps due to the sporulation of the microbial community (Gao et al., 2011). The system was successfully reached a steady state with a significant change in the total biogas production to 0.4 L/L/day and gas composition of 45.6% and 54.4% hydrogen and carbon dioxide gases, respectively. At the second shock, a decrease in the biogas production from 0.4 to 0.18 L/L/ day occurred with a 45–29% shift of hydrogen gas. The biogas production and hydrogen gas percentage were successfully recovered to the previous steady state condition after the second shock with temperature recovery (Iranpour et al., 2005). It appears that the fermentation temperature and temperature shock have fundamental impacts on the microbial community function. Gao et al. (2011) reported Clostridium species were dominant at 37–45 °C while at 50–55 °C and 60 °C the culture was dominated by Bacillus coagulans and Thermoanaerobacterium, respectively, consequently a change in environment temperature would lead to changes in microbial community function (Tang et al., 2008). Our results show that the changing in the reactor performance before and after the shock was maybe due to changes in the microbial community function coupled with changing in soluble by-products in this study.

3.1.3. Hyper-thermophilic temperature The time course of the biogas production rates, H2 concentrations, pH, VFA and VSS in the hyper-thermophilic reactor are presented in Fig. 4. The system was continuously operated for five months at 80 ± 1 °C to investigate effect of temperature and

temperature shock. Throughout the steady state condition, the maximum biogas production was 0.47 L/L/day with 46.6% and 53.4% hydrogen and carbon dioxide gases, respectively. This result confirmed that the thermophilic temperature or higher was able to inhibit methane generation during cellulose fermentation by anaerobic mixed microflora. A pH of 5.98 ± 0.05 was maintained naturally during the experiment with the use sodium bicarbonate. Under this condition, the system was exposed to a 44 °C temperature shock from 80 to 36 ± 2 °C, resulting in a drop in biogas production to 0.23 L/L/day with 23% and 77% hydrogen and carbon dioxide gases, respectively. The system was successfully recovered to a steady state with considerable drop in biogas production to 0.35 L/L/day and 43% hydrogen gas. At the second shock, the biogas production decreased to 0.2 L/L/day with 21% hydrogen gas. The performance of the system successfully recovered to the previous steady state condition with insignificant change from the reading from the first temperature shock. The biogas production reached 0.36 L/L/day with 43% and 57% hydrogen and carbon dioxide gases, respectively. Our results agreed with other previous study reported that the hyper-thermophilic mixed microflora was able to resume growth after temperature shock (Boonyaratanakornkit et al., 2005) and the thermophilic and hyper-thermophilic microorganisms were more resilient to temperature shock than mesophilic mixed microflora to recover hydrogen gas production. Gao et al. (2011) reported that under hyper-thermophilic temperature, the microbial community had low diversity comparing with mesophilic and thermophilic temperature. Therefore, low pathogens and hydrogen consumer bacteria in digested residues were achieved compare with mesophilic temperature. In addition, the biohydrogen reaction at high temperature was favorable and made dark hydrogen fermentation more energetic while the hydrogen utilization processes were negatively affected with temperature increase (Amend and shock, 2001).

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3.2. Effect of temperature and temperature shock on the metabolic stoichiometry of cellulose fermentation 3.2.1. Bio-hydrogen yield Under the steady-state conditions, the metabolic products are compatible, and there is no accumulation of the end products in the system (Peck et al., 1986). However, the unexpected changes in the operating temperature results unbalanced in the fermentation process (Temper et al., 1983). This was observed in our three working reactors. According to metabolic products of cellulose fermentation in this study, the stoichiometry equations of the hydrogen fermentation of cellulose under different fermentation temperature during the steady state condition and temperature shock were calculated as follows:[The mesophilic reactor]

ðC6 H10 O5 Þn ! 0:97nCH4 þ 0:62nH2 þ 0:59nC2 H4 O2 þ 0:32nC4 H7 O2 þ 0:12nC3 H6 O2n þ 0:03nC2 H6 O ðC6 H10 O5 Þn

1st temp:shock

!

ð1Þ

0:77nCH4 þ 0:55nH2 þ 0:65nC2 H4 O2

þ 0:24nC4 H7 O2 þ 0:13nC3 H6 O2 þ 0:03nC2 H6 O ðC6 H10 O5 Þn

2nd temp:shock

!

ð2Þ

0:87nCH4 þ 0:16nH2 þ 0:53nC2 H4 O2

þ 0:19nC4 H7 O2 þ 0:21nC3 H6 O2 þ 0:01nC2 H6 O

ð3Þ

[The thermophilic reactor]

ðC6 H10 O5 Þn ! 2:21nH2 þ 0:71nC2 H4 O2 þ 0:33nC4 H7 O2 þ 0:05nC2 H6 O ðC6 H10 O5 Þn

1st temp shock

!

1:47nH2 þ 0:52nC2 H4 O2

2nd temp shock

!

1st temp shock

!

1:21nH2 þ 0:43nC2 H4 O2

þ 0:11nC4 H7 O2 þ 0:17nC2 H6 O ðC6 H10 O5 Þn

2nd temp shock

!

ð8Þ

1:18nH2 þ 0:41nC2 H4 O2

þ 0:13nC4 H7 O2 þ 0:14nC2 H6 O

ð9Þ

Eqs. (1, 4, and 7) show the metabolic products of cellulose fermentation during the steady state condition under various temperatures before the shock. The fermentative characteristics in each reactor, including the yields of hydrogen and the intermediate products clearly differed at various fermentation temperatures. The deference in hydrogen yield, at 0.64–2.21 mol/mol hexose, under mesophilic and thermophilic temperatures, respectively, may be attributed to the inhibition of methanogenic and hydrogen consuming bacteria (Spyros et al., 1990 and Lay, 2001). At hyperthermophilic temperature, hydrogen yield reached 1.75 mol/mol hexose. The results showed that the thermophilic fermentation is still expected having a better economic performance for bio-hydrogen production. With regard to temperature shock, the change in the hydrogen yield after temperature shock in all reactors is given by Eqs. (2, 3, 5, 6, 8, and 9). Under the thermophilic and hyperthermophilic temperatures, insignificant change was noted in the hydrogen yield after the second shock despite the significant slowing of the hydrogen yield (2.21–1.47 and 1.75–1.21 mol/mol hexose) in both reactors after the first shock. On the other hand, the detrimental effect of the second shock on the mesophilic reactor performance in terms hydrogen yield (0.16 mol/mol hexose); however, methane yield was successfully recovered and maintained at 0.87 mol/mol hexose.

ð4Þ

þ 0:25nC4 H7 O2 þ 0:12nC2 H6 O ðC6 H10 O5 Þn

ðC6 H10 O5 Þn

ð5Þ

1:48nH2 þ 0:57nC2 H4 O2

þ 0:27nC4 H7 O2 þ 0:09nC2 H6 O

ð6Þ

[The hyper-thermophilic reactor]

ðC6 H10 O5 Þn ! 1:75nH2 þ 0:58nC2 H4 O2 þ 0:14nC4 H7 O2 þ 0:06nC2 H6 O

ð7Þ

3.2.2. Liquid end-products The liquid end-product results of cellulose fermentation under different fermentation temperatures and temperature shocks are shown in Eqs. ((1)–(9)) and Fig. 5. All measurements reported were taken during the steady state condition. During the cellulose fermentation, acetate, butyrate, together with ethanol were the main by-products in the liquid phase in the thermophilic and hyperthermophilic fermentation temperature; as well as propionic acid was produced in the mesophilic condition has been observed in other reports (Liu et al., 2003 and Lay, 2001). The acetate, butyrate and ethanol reached 0.62, 0.33, 0.05 and 0.68, 0.14, 0.06 mol/mol hexose under thermophilic and hyper-thermophilic conditions,

0.8 Acetic acid Butyric Acid

0.7

Propoinic Acid Ethanol

Mol/mol hexose

0.6 0.5 0.4 0.3 0.2 0.1 0

Mesophilic reactor

Thermophilic reactor

Hyper-thermophilic reactor

Fig. 5. Effect of temperature and temperature shock on the by-products production.

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100 90 80 70

COD (%)

60 Methane

50

Hydrogen

40

by-products

30

Cellulose

20 10 0 Cellulose

before Shock

1st Shock

2nd Shock

Mesophilic Reactor

before Shock

1st Shock

2nd Shock

Thermophilic Reactor

before Shock

1st Shock

2nd Shock

Hyperthermophilic Reactor

Fig. 6. Effect of temperature and temperature shock on the COD mass balances.

respectively. The results suggest that the digester sludge under thermophilic and hyper-thermophilic temperatures possessed clostridial characteristics in the transformation of the cellulose to hydrogen as evidenced by the typical hydrogen-/acids- and alcohols-producing phases (Payot et al., 1998 and Carver et al., 2012). Under mesophilic condition, as well as acetate- butyrate fermentation, together with ethanol was observed the propionic acid type also was detected; these by-products concentration reached 0.59, 0.32, 0.12 and 0.03 mol/mol hexose, respectively. The results showed the thermophilic and hyper-thermophilic temperatures could effectively enhance hydrogen production from cellulose comparing with mesophilic temperature (Lin and Hung, 2008). These results parallel those with bacterial mixed culture from sewage sludge fermenting cellulose to acetic acid, propionic acid, H2 and CO2, with ethanol produced in smaller amounts (Khan et al., 1981). In respect of temperature shock, a significant change in the soluble by-products after the shock was noted in all reactors (see Fig. 5). Under mesophilic condition, the acetate and propionic acid concentrations increased 10% and 8%, respectively; however butyrate production decreased by 25% and no change was detected in ethanol production after the first shock. The propionic acid increased by 61%; however, acetate, butyrate and ethanol production decreased by 18.5%, 20.8% and 67%, respectively after the second shock. The possible utilization of hydrogen in propionate formation makes this process unfavorable to hydrogen production The acetate and butyrate concentrations were decreased by 16%, 26% and 24%, 21%; while, ethanol production was increased by 140% and 183% under thermophilic and hyper-thermophilic conditions, respectively after the first shock. The above findings indicate that the microbial community was able to induce a metabolic shift from hydrogen VFAs to alcohols; suggested that the digester sludge possessed Moorella sp characteristics in the transformation of the hydrogen and CO2 to ethanol production at pH 5 (Sakai et al., 2004). The acetate and butyrate production were increased by 8% and 9.6% under thermophilic condition, respectively; while, the butyrate production increased by 18% and the acetate decreased by 4.7% under hyper-thermophilic conditions after the second shock. At the same time, the ethanol production decreased by 25% and 23.5%, respectively. These results suggested that a change in environment temperature would lead to changes in microbial community function and the thermophilic and hyper-thermophilic temperature seems to be more reliable after the first shock compare with mesophilic temperature.

3.3. System recovery period The recovery period can be defined as ‘‘A period between two different steady state conditions, where significant changes in the reactor performance are caused by removing the temporary effect, and followed by recovery of the microflora activity’’. All reactors were subjected to two temperature shock during the steady state condition. The thermophilic and hyper-thermophilic reactors were significant affected by first temperature shock; however insignificant change was detected after the second shock (see Figs. 3 and 4). On the other hand, the detrimental effect of the second shock on the system performance in terms of hydrogen gas was significant under mesophilic temperature. After the first shock, the mesophilic, thermophilic and hyper-thermophilic reactors were reached steady state condition after 8, 12 and 16 days, respectively. That means the recovery period was increased by increasing of the fermentation temperature. This may result because the spores were transferred completely into a fully active vegetative cell requires a lag period for most spore-forming species (Talaro and Talaro, 1993 and Iranpour et al., 2005). Another important consideration is the function of the microbial community after first shock. In the mesophilic reactor, the performance was not significantly affected by temperature shock with regard to either gas production or composition. On the other hand, a significant downward shift in the performance of thermophilic and hyper-thermophilic reactors in biological hydrogen production was noted at 34.5% and 31% respectively. With regard to the second shock, the thermophilic and hyper-thermophilic reactors were successfully recovered to the previous steady state condition within 8 and 12 days respectively. While the mesophilic reactor recovered within 8 days, which is the same recovery period in the first shock. Ahn and Forster (2002) reported that the mesophilic digester (35 °C) was restored to the initial steady state within 8 days after 11– 17 °C temperature shock. Our results show that the recovery period after the second shock was shorter than after the first shock under thermophilic and hyper-thermophilic reactors. In addition, the performance of the thermophilic and hyper-thermophilic reactors completely recovered to the steady state after second temperature shock (see Figs. 3 and 4). This implies that the microbial community structure changed to adapt such a condition. On the other hand, hydrogen production did not recover in the mesophilic reactor (Fig. 2) and hydrogen production was significantly reduced 68%; but methane production recovered in this system (Peck

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Table 1 Effect of temperature and temperature shock on reactors performance under steady state condition. pH

37 ± 1 °C 37 ± 1 °C 37 ± 1 °C 55 ± 1 °C 55 ± 1 °C 55 ± 1 °C 80 ± 1 °C 80 ± 1 °C 80 ± 1 °C

before temp. shock after 1st temp. shock after 2nd temp. shock before temp. shock after 1st temp. shock after 2nd temp. shock before temp. shock after 1st temp. shock after 2nd temp. shock

5.59 ± 0.2 5.63 ± 0.1 5.62 ± 0.1 5.86 ± 0.1 5.88 ± 0.2 5.81 ± 0.2 5.98 ± 0.1 6.01 ± 0.1 6.05 ± 0.1

Hydrogen yield mmol/g cellulose

3.6 ± 0.1 3.1 ± 0.1 0.9 ± 0. 2 12.3 ± 0.6 8.1 ± 0.4 8.2 ± 0.3 9.7 ± 0.4 6.7 ± 0.4 6.6 ± 0.5

Methane yield mmol/g cellulose

5.4 ± 0.26 4.2 ± 0.19 4.8 ± 0.71 – – – – – –

et al., 1986). This suggested that the thermal pretreatment for digested sewage sludge was recommended to generate hydrogen from cellulose by mesophilic temperature (Lay, 2001). 3.4. The effect of temperature and temperature shock on COD removal and mass balance The COD mass balance was calculated according to the COD conversion coefficient of each product. Fig. 6 illustrates the influence of different temperatures and temperature shock on COD mass balance under the steady state condition. The results show that the maximum cellulose degradation reached 71.3% at mesophilic temperature; however, it decreased to 60.3% and 53% under thermophilic and hyper-thermophilic temperature, respectively (see Table 1). While, the maximum hydrogen yield obtained by thermophilic temperature. The cellulose degradation decreased by temperature shock to 68.7, 51.2 and 44.9 under mesophilic, thermophilic and hyper-thermophilic temperature. No significant change was detected in cellulose degradation under thermophilic and hyper-thermophilic after the second shock. The experimental results show that the optimum performance was obtained at the thermophilic temperature (55 °C) due to high cellulose degradation, high hydrogen production rates and stability after temperature shock. Our results are in complete agreement with those reported by Liu et al. (2003) who found that thermophilic treatment (55 °C) was more effective in the treatment of cellulosecontaining wastewater without prior treatment than mesophilic treatment. In contrast, high methane and low hydrogen production produced under mesophilic temperature were obtained (Lay, 2001).

4. Conclusion The 55 ± 1 and 80 ± 1 °C presented stable cellulosic- hydrogen fermentation, while the 37 ± 1 °C presented a lower performance as well as methane production. The hydrogen production by 55 ± 1 °C is considered a sustainable way to produce renewable hydrogen compared with 37 ± 1 and 80 ± 1 °C. Under 55 ± 1 and 80 ± 1 °C, the hydrogen yield was negatively affected by the first shock and insignificant changes were detected after the second shock. Whereas, a very low hydrogen yield was maintained after the second shock, the methane yield was increased under 37 ± 1 °C. The reactor performance under 55 ± 1 and 80 ± 1 °C appeared to be more resilient to the temperature decreases than 37 ± 1 °C. Acknowledgement This work is partially supported by the Government of Egypt.

Major soluble by-products VFA (g/L)

Ethanol (g/L)

Acetic A

Butyric A

Propionic A

1.97 2.16 1.74 2.36 1.71 1.90 1.93 1.43 1.40

1.53 1.13 0.93 1.61 1.22 1.32 0.67 0.52 0.61

0.49 0.52 0.86 – – – – – –

0.077 0.074 0.026 0.13 0.31 0.23 0.16 0.43 0.36

Cellulose Degradation (%)

71.3 68.7 65.1 60.3 51.2 50.7 53.3 44.9 44.6

References APHA, 2005. Standard Methods for the Examination of Water and Wastewater. American Public Health Association, New York. Ahn, J.-H., Forster, C.F., 2002. The effect of temperature variations on the performance of mesophilic and thermophilic anaerobic filters treating a simulated papermill wastewater. Process Biochem. 37, 589–594. Ahring, B.K., Ibrahim, A.A., Mladenovska, Z., 2001. Effect of temperature increase from 55 to 65 °C on performance and microbial population dynamics of an anaerobic reactor treating cattle manure. Water Res. 35, 2446–2452. Amend, J.P., Shock, E.L., 2001. Energetics of overall metabolic reactions of thermophilic and hyperthermophilic archaea and bacteria. FEMS Microbiol. Rev. 25 (2), 175–243. Angenent, L.T., Abel, S.J., Sung, S., 2002. Effect of an organic shock load on the stability of an anaerobic migrating blanket reactor. J. Environ. Eng. 128 (12), 1109–1120. Berg, L., van den, Lentz, C. P., 1981. Performance and stability of the anaerobic contact process as affected by waste composition, inoculation and solids retention time. In: Proc. 35th Ind. Waste Conf. Purdue University, pp. 496–501 Boonyaratanakornkit, B.B., Simpson, A.J., Whitehead, T.A., Fraser, C.M., El-Sayed, N.M., Clark, D.S., 2005. Transcriptional profiling of the hyperthermophilic methanarchaeon Methanococcus jannaschii in response to lethal heat and nonlethal cold shock. Environ. Microbiol. 7 (6), 789–797. Carver, S.M., Nelson, M.C., Lepistö, R., Yu, Z., Tuovinen, O.H., 2012. Hydrogen and volatile fatty acid production during fermentation of cellulosic substrates by a thermophilic consortium at 50 and 60 °C. Bioresour. Technol. 104, 424–431. Ewan, B.C.R., Allen, R.W.K., 2005. A figure of merit assessment of the routes to hydrogen. Int. J. Hydrogen Energy 30, 809–819. Gadow, S.I., Li, Y.Y., Liu, Y., 2012. Effect of temperature on continuous hydrogen production of cellulose. Int. J. Hydrogen Energy 37, 15465–15472. Gao, W.J., Leung, K.T., Qin, W.S., Liao, B.Q., 2011. Effects of temperature and temperature shock on the performance and microbial community structure of a submerged anaerobic membrane bioreactor. Bioresour. Technol. 102, 8733– 8740. Graef, S.P., Andrews, J.F., 1974. Stability and control of anaerobic digestion. J. Water Pollut Control Fed. 46, 666–683. Iranpour, R., Alatriste-Mondragon, F., Cox, H.H., Haug, R.T., 2005. Effects of transient temperature increases on odor production from thermophilic anaerobic digestion. Water Sci. Technol. 52, 229–235. Khan, A.W., Duncan, W.L., Van, D.B., 1981. Fermentative conversion of cellulose to acetic acid and cellulolytic enzyme production by a bacterial mixed culture obtained from sewage sludge. Appl. Environ. Microbiol., 1214–1218. Kumar, G., Lin, C.Y., 2013. Bioconversion of de-oiled Jatropha Waste (DJW) to hydrogen and methane gas by anaerobic fermentation: influence of substrate concentration, temperature and pH. Int. J. Hydrogen Energy 38 (1), 63–72. Lay, J.J., 2001. Biohydrogen generation by mesophilic anaerobic fermentation of microcrystalline cellulose. Biotechnol. Bioeng. 74, 281–287. Lee, D.Y., Ebie, Y., Xu, K.Q., Li, Y.Y., Inamori, Y., 2010. Continuous H2 and CH4 production from high-solid food waste in the two-stage thermophilic fermentation process with the recirculation of digester sludge. Bioresour. Technol. 101, S42–S47. Lin, C.Y., Hung, W.C., 2008. Enhancement of fermentative hydrogen/ethanol production from cellulose using mixed anaerobic cultures. Int. J. Hydrogen Energy 33, 3660–3667. Liu, H., Zhang, T., Fang Herbert, H.P., 2003. Thermophilic H2 production from a cellulose-containing wastewater. Biotechnol. Lett. 25, 365–369. Luo, G., Karakashev, D., Xie, L., Zhou, Q., 2011. Angelidaki I. Long-term effect of inoculum pretreatment on fermentative hydrogen production by repeated batch cultivations: homoacetogenesis and methanogenesis as competitors to hydrogen production. Biotechnol. Bioeng. 108 (8), 1816–1827. Lynd, L.R., Weimer, P.J., Zyl, W.H., Pretorius, I.S., 2002. Microbial cellulose utilization: fundamentals and biotechnology. Microbiol. Mol. Biol. Rev. 66, 506–577. Obaya, M.C., Valdes, E., Ramos, J., 1994. Stability studies of thermophilic anaerobic sludges under suboptimal feeding conditions and temperatures. Acta Biotechnol. 14 (2), 193–198.

S.I Gadow et al. / Bioresource Technology 142 (2013) 304–311 Payot, S., Guedon, E., Cailliez, C., Gelhage, E., Petitdemange, H., 1998. Metabolism of cellobiose by Clostridium cellulolyticum growing in continuous culture: evidence for decreased NADH reoxidation as a factor limiting growth. Microbiology 144, 375–384. Peck, M.W., Skilton, J.M., Hawkes, F.R., Hawkes, D.L., 1986. Effects of temperature shock treatments on the stability of anaerobic digesters operated on separated cattle slurry. Water Res. 20 (4), 453–462. Sahlstrom, L., 2003. A review of survival of pathogenic bacteria in organic waste used in biogas plants. Bioresour. Technol. 87 (2), 161–166. Sakai, S., Nakashimada, Y., Yoshimoto, H., Watanabe, S., Okada, H., Nishio, N., 2004. Ethanol production from H2 and CO2 by a newly isolated thermophilic bacterium, Moorella sp. HUC22-1. Biotechnol. Lett. 20, 1607–1612. Speece, R.E., 1983. Anaerobic biotechnology for the industrial wastewater treatment. Environ. Sci. Technol. 17, 416A–427A. Spyros, G.P., Terry, L.M., Meyer, J.W., 1990. Cellulose fermentation by continuous cultures of Ruminococcus albus and Methanobrevibacter smithii. Appl. Microbiol. Biotechnol. 33, 109–116.

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Talaro, K., Talaro, A., 1993. Foundations in Microbiology. Wm. C.Brown, Dubuque, IA. Tang, Y.Q., Matsui, T., Morimura, S., Wu, X.L., Kida, K., 2008. Effect of temperature on microbial community of a glucose-degrading methanogenic consortium under hyperthermophilic chemostat cultivation. J. Biosci. Bioeng. 106 (2), 180–187. Temper, U., Winter, J., Kandler, O., 1983. Methane fermentation of wastes at mesophilic and thermophilic temperatures. In: Straub, A., Chartier, P. and Schleser, G. (Eds.), Energy from Biomass, Appl. Sci. London. 521–525 Van Lier, J.B., Rintala, J., Sanz Martin, J.L., Lettinga, G., 1990. Effect of short-term temperature increase on the performance of a mesophilic UASB reactor. Water Sci. Technol. 22 (9), 183–190. Van Niel, E.W.J., Budde, M.A.W., de Haas, G.G., van der Wa, F.J., Claasen, P.A.M., St ams, A.J.M., 2002. Distinctive properties of high hydrogen producing extreme thermophiles, Caldicellulosiruptor saccharolyticus and Thermotoga elfii. Int. J. Hydrogen Energy 27 (11–12), 1391–1398. Zhao, Q.B., Yu, H.Q., 2008. Fermentative H2 production in an upflow anaerobic sludge blanket reactor at various pH values. Bioresour. Technol. 99 (5), 1353–1358.