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Effect of heat pre-treatment temperature on isolation of hydrogen producing functional consortium from soil
Abstract
A functional hydrogen producing consortium was isolated from soil by heat pre-treatment technique and hydrogen production at different substrate concentration was evaluated. The forest soil was heat pre-treated at 65, 80, 95, 105 and 120 °C temperature for 1 h. As revealed by PCR-DGGE analysis and hydrogen yield, the hydrogen producing microbial community changed with increase in heat pre-treatment temperatures giving potential hydrogen producing consortium at 95–105 °C soil pre-treatment. The maximum hydrogen production rate, hydrogen yield and cumulative hydrogen with 15–20 g glucose were 1390–1576 mL/L/day, 1.83–1.93 mol H2/mol glucose, and 2966–3146 mL H2/L, respectively. The metabolic pathways shifted from ethanol-type to acetate–formate type as soil pre-treatment temperature increased from 65 to 120 °C. The soil heat pre-treatment approach is effective for isolating hydrogen producing natural Clostridium consortium from the soil as enumerations of the functional strains need specific temperature range to florish.
... The heat pretreatment method on the mesophilic sludge inactivates the hydrogenotrophic methanogens, improves the availability of hydrogen producing microorganisms and produces more hydrogen in comparison to no pretreatment (control production) [41]. The heat-treatment temperatures use varied from 65 °C to 121 °C and the duration varied from 10 min to 24 h [29,34,35,46]. Statistical models to determine the optimum temperature and heating time for seed inoculum such as mountain soil have been developed [46]. ...
... The heat-treatment temperatures use varied from 65 °C to 121 °C and the duration varied from 10 min to 24 h [29,34,35,46]. Statistical models to determine the optimum temperature and heating time for seed inoculum such as mountain soil have been developed [46]. The optimized conditions of 105 °C for 30 min resulted in an increased hydrogen yield of 318 mL H2/g of solid or 1.93 mol H2/mol glucose in comparison to untreated soil (control production) of 40-60 mL H2/g of solid [46] as summarized in Table 1. ...
... Statistical models to determine the optimum temperature and heating time for seed inoculum such as mountain soil have been developed [46]. The optimized conditions of 105 °C for 30 min resulted in an increased hydrogen yield of 318 mL H2/g of solid or 1.93 mol H2/mol glucose in comparison to untreated soil (control production) of 40-60 mL H2/g of solid [46] as summarized in Table 1. The heat-treatment plays a very important role in the metabolic pathway, with an increase in temperature from 65 to 120 °C resulting in a pathway shift from ethanol-type to acetate-formate type for hydrogen production. ...
Hydrogen is an important source of energy and is considered as the future energy carrier post-petroleum era. Nowadays hydrogen production through various methods is being explored and developed to minimize the production costs. Biological hydrogen production has remained an attractive option, highly economical despite low yields. The mixed-culture systems use undefined microbial consortia unlike pure-cultures that use defined microbial species for hydrogen production. This review summarizes mixed-culture system pretreatments such as heat, chemical (acid, alkali), microwave, ultrasound, aeration, and electric current, amongst others, and their combinations to improve the hydrogen yields. The literature representation of pretreatments in mixed-culture systems is as follows: 45-50% heat-treatment, 15-20% chemical, 5-10% microwave, 10-15% combined and 10-15% other treatment. In comparison to pure-culture mixed-culture offers several advantages, such as technical feasibility, minimum inoculum steps, minimum media supplements, ease of operation, and the fact it works on a wide spectrum of low-cost easily available organic wastes for valorization in hydrogen production. In comparison to pure-culture, mixed-culture can eliminate media sterilization (4 h), incubation step (18-36 h), media supplements cost ($4-6 for bioconversion of 1 kg crude glycerol (CG)) and around 10-15 Millijoule (MJ) of energy can be decreased for the single run.
... The heat pretreatment method on the mesophilic sludge inactivates the hydrogenotrophic methanogens, improves the availability of hydrogen producing microorganisms and produces more hydrogen in comparison to no pretreatment (control production) [41]. The heat-treatment temperatures use varied from 65 °C to 121 °C and the duration varied from 10 min to 24 h [29,34,35,46]. Statistical models to determine the optimum temperature and heating time for seed inoculum such as mountain soil have been developed [46]. ...
... The heat-treatment temperatures use varied from 65 °C to 121 °C and the duration varied from 10 min to 24 h [29,34,35,46]. Statistical models to determine the optimum temperature and heating time for seed inoculum such as mountain soil have been developed [46]. The optimized conditions of 105 °C for 30 min resulted in an increased hydrogen yield of 318 mL H2/g of solid or 1.93 mol H2/mol glucose in comparison to untreated soil (control production) of 40-60 mL H2/g of solid [46] as summarized in Table 1. ...
... Statistical models to determine the optimum temperature and heating time for seed inoculum such as mountain soil have been developed [46]. The optimized conditions of 105 °C for 30 min resulted in an increased hydrogen yield of 318 mL H2/g of solid or 1.93 mol H2/mol glucose in comparison to untreated soil (control production) of 40-60 mL H2/g of solid [46] as summarized in Table 1. The heat-treatment plays a very important role in the metabolic pathway, with an increase in temperature from 65 to 120 °C resulting in a pathway shift from ethanol-type to acetate-formate type for hydrogen production. ...
Hydrogen is an important source of energy and is considered as the future energy carrier post-petroleum era. Nowadays hydrogen production through various methods is being explored and developed to minimize the production costs. Biological hydrogen production has remained an attractive option, highly economical despite low yields. The mixed-culture systems use undefined microbial consortia unlike pure-cultures that use defined microbial species for hydrogen production. This review summarizes mixed-culture system pretreatments such as heat, chemical (acid, alkali), microwave, ultrasound, aeration, and electric current, amongst others, and their combinations to improve the hydrogen yields. The literature representation of pretreatments in mixed-culture systems is as follows: 45–50% heat-treatment, 15–20% chemical, 5–10% microwave, 10–15% combined and 10–15% other treatment. In comparison to pure-culture mixed-culture offers several advantages, such as technical feasibility, minimum inoculum steps, minimum media supplements, ease of operation, and the fact it works on a wide spectrum of low-cost easily available organic wastes for valorization in hydrogen production. In comparison to pure-culture, mixed-culture can eliminate media sterilization (4 h), incubation step (18–36 h), media supplements cost ($4–6 for bioconversion of 1 kg crude glycerol (CG)) and around 10–15 Millijoule (MJ) of energy can be decreased for the single run.
... Therefore, further studies are neces- sary to clarify the best conditions. In this study, we compared the effects of different heat pretreatment temperatures (50, 60, 70, 80, 90, 100, and 110 C) and time periods (10,20,30,40,50,60,90, and 120 min) on the efficiency of hydrogen-producing bacteria enrichment. We investigated the effects of heat pretreatment on the perfor- mance of the hydrogen production process and the microbial communities. ...
... In the heat pretreatment test, the sludge samples were heated at temperatures of 50, 60, 70, 80, 90, 100, and 110 C for 30 min to enrich the hydrogen-producing inocula. In the heat pretreatment test for different time pe- riods, the sludge samples were heated for 10,20,30,40,50,60,90, and 120 min at 70 C to enrich the hydrogen-producing inocula. ...
... Wong et al. [20] found that heat pretreatment at 65 C pro- duced the highest hydrogen yield among the different pre- treatment temperatures that they tested (40, 55, 65, 80 and 95 C), where further increases in the temperature decreased the hydrogen yield. Ravindran et al. [29] found that 95 C was the most effective pretreatment temperature for hydrogen production, where they compared temperatures of 65, 80, 95, 105, and 120 C [30]. In another study, incubated sewage sludge was pretreated at 80 C for 30 min [31]. ...
Potential of pH decrease is one of the major obstacle in stable operation in coculture of dark- and photo-fermentative bacteria for hydrogen production. In this study, a dark fermentative bacterial consort and acid-tolerant marine photo-fermentative bacterium, Rhodovulum sulfidophilum TH-102, were individual or co-cultured in high salt medium for hydrogen production. All co-cultures produced more hydrogen than the individual culture of photo or dark fermentation. The dark/photo bacterial ratios were 1:5, 1:10, 1:15 and 1:20, respectively. Among the coculture ratios, bacterial ratio 1:10 produced the highest hydrogen yield (1694 ± 21 mL/L). The addition of the photo-fermentative bacterium to the dark fermentation consort stabilized the pH value and decreased the oxidation-reduction potential of the co-culture system and extended the hydrogen production period. Carbon fixation by the photo-fermentative bacterium may play some role in improving the hydrogen yield of the co-culture system.
... In this study, we compared the effects of different heat pretreatment temperatures (50, 60, 70, 80, 90, 100, and 110 C) and time periods (10,20,30,40,50,60,90, and 120 min) on the efficiency of hydrogen-producing bacteria enrichment. We investigated the effects of heat pretreatment on the performance of the hydrogen production process and the microbial communities. ...
... Wong et al. [20] found that heat pretreatment at 65 C produced the highest hydrogen yield among the different pretreatment temperatures that they tested (40, 55, 65, 80 and 95 C), where further increases in the temperature decreased the hydrogen yield. Ravindran et al. [29] found that 95 C was the most effective pretreatment temperature for hydrogen production, where they compared temperatures of 65, 80, 95, 105, and 120 C [30]. In another study, incubated sewage sludge was pretreated at 80 C for 30 min [31]. ...
... The control test contained the most bands. This suggests that the heat pretreatment reduced the diversity of species, as shown in previous studies [30]. Homoacetogens (Acetobacterium Woodii, band 1 and 6) are hydrogen-consuming bacteria and they have inhibitory effects on other hydrogenproducing microorganisms [14], and they were only found in the control and low temperature (50 C) treatment cultures. ...
Salt-containing wastes have increased in abundance in recent years. In this study, we investigated hydrogen production from intensive shrimp mariculture organic waste in batch culture experiments. Sludge samples were pretreated at different temperatures (50–110 °C) and for various time periods (10–120 min) to enrich the hydrogen-producing microflora. The results showed that all of the thermal pretreatments achieved higher hydrogen yields compared with those obtained in the experiments without heat treatment. Pretreatment at 70 °C achieved the maximum hydrogen yield. Different heat pretreatment time periods between 30 and 120 min had little influence on the hydrogen yield. Considering the energy consumption of the pretreatment process, 30 min at 70 °C were selected as the optimum pretreatment conditions. Microbial community DNA analysis and the diversity of the hydA gene showed that the number of bacterial species and the hydA gene diversity decreased as the pretreatment temperature increased.
... Only a few studies analyzed the effect of heat pretreatment at different temperatures on mesophilic hydrogen production (Baghchehsaraee et al. 2008;Ravindran et al. 2010). Baghchehsaraee et al. (2008) produced hydrogen from glucose with anaerobically digested sludge pretreated at 65, 80 or 95°C. ...
... The bacterial community analysis showed that elevated temperatures reduced species diversity. On the other hand, Ravindran et al. (2010) in experiments with inoculum (forest soil) pretreated at 65, 80, 95, 105, and 120°C reported the highest H 2 production for 105°C (1.92 mol H 2 /mol glucose ). When inoculum was pretreated at 105°C, the presence of additional bacterial species was detected. ...
Hydrogen produced from lignocellulose biomass is deemed as a promising fuel of the future. However, direct cellulose utilization remains an issue due to the low hydrogen yields. In this study, the long-term effect of inoculum (anaerobic sludge) heat pretreatment on hydrogen production from untreated cellulose and starch was evaluated during repeated batch processes. The inoculum pretreatment at 90°C was not sufficient to suppress H2 consuming bacteria, both for starch and cellulose. Although hydrogen was produced, it was rapidly utilized with simultaneous accumulation of acetic and propionic acid. The pretreatment at 100°C (20 min) resulted in the successful enrichment of hydrogen producers on starch. High production of hydrogen (1.2 l H2/lmedium) and H2 yield (1.7 mol H2/molhexose) were maintained for 130 days, with butyric (1.5 g/l) and acetic acid (0.65 g/l) as main byproducts. On the other hand, the process with cellulose showed lower hydrogen production (0.3 l H2/lmedium) with simultaneous high acetic acid (1.4 g/l) and ethanol (1.2 g/l) concentration. Elimination of sulfates from the medium led to the efficient production of hydrogen in the initial cycles – 0.97 mol H2/molhexose (5.93 mmol H2/gcellulose). However, the effectiveness of pretreatment was only temporary for cellulose, because propionic acid accumulation (1.5 g/l) was observed after 25 days, which resulted in lower H2 production. The effective production of hydrogen from cellulose was also maintained for 40 days in a repeated fed-batch process (0.63 mol H2/molhexose).
... In addition, substrate pretreatment, and pretreatment of the microbial seed culture, have been found to be useful for effective acidogenesis and VFA production. To achieve competent microbial communities for hydrogen production and generation of VFAs, different pretreatment methods for the anaerobic inoculum, such as heat shock, chemical application, aeration, microwaves, and ultrasound have been thoroughly investigated [15,16]. Not only are process conditions, pretreatment methods, microbial structure and metabolism important in the production of VHAs, but also their concentration and chemical composition ( Figure 4). ...
Global energy consumption has been increasing in tandem with economic growth motivating researchers to focus on renewable energy sources. Dark fermentative hydrogen synthesis utilizing various biomass resources is a promising, less costly, and less energy-intensive bioprocess relative to other biohydrogen production routes. The generated acidogenic dark fermentative effluent [e.g., volatile fatty acids (VFAs)] has potential as a reliable and sustainable carbon substrate for polyhydroxyalkanoate (PHA) synthesis. PHA, an important alternative to petrochemical based polymers has attracted interest recently, owing to its biodegradability and biocompatibility. This review illustrates methods for the conversion of acidogenic effluents (VFAs), such as acetate, butyrate, propionate, lactate, valerate, and mixtures of VFAs, into the value-added compound PHA. In addition, the review provides a comprehensive update on research progress of VFAs to PHA conversion and related enhancement techniques including optimization of operational parameters, fermentation strategies, and genetic engineering approaches. Finally, potential bottlenecks and future directions for the conversion of VFAs to PHA are outlined. This review offers insights to researchers on an integrated biorefinery route for sustainable and cost-effective bioplastics production.
... This concurred well with previous work which revealed that acid treatment of sludge resulted in decreased H 2 production [42]. Previous studies have reported that because H 2producing cultures are spore-forming bacteria, their capacity to resist heat shock or other severe pretreatment enables i n t e r n a t i o n a l j o u r n a l o f h y d r o g e n e n e r g y x x x ( x x x x ) x x x them to be easily and efficiently selected from mixed inoculum, with long-term stability [43,44]. ...
Methods are investigated to prepare active hydrogen (H2)-producing cultures originating from palm oil mill sludge using dark fermentation. The first successful method that produces potent H2-producing cultures and avoids growing H2-consuming methanogens involves heat pretreatment of the sludge at 100 °C for 2 h and then the sludge sample is shocked in an ice bath for 15 min. Subsequently, a glucose solution rich in nutrients (glucose-based substrate) of 14.80 g chemical oxygen demand (COD)/L is fed in to enrich the H2-producing cultures. The H2 production reaches 78.63% on day 31. The second method involves acid pretreatment of sludge with 10 M hydrochloric acid at pH 3 for 48 h. Glucose-based substrate of 25.47 g COD/L is fed into the system. The H2 production is 69.41% on day 27. For both methods, the H2 production is stable after the H2 content reached its maximum. The operation is performed semi-continuously using a hydraulic retention time of 1 day and at 30 °C. The optimum bacterial cells-to-COD level of substrate is approximately 0.60 in the start-up medium. The fermentation medium has an optimum initial pH of 5 and a final pH of 5.2–5.3. These two methods are recommended to produce active H2-producing cultures for plant start-up in bio-H2 production.
... Heat treatments are also used to isolate functional consortia for H 2 production. Ravindran et al. (2010) isolated a bacterial consortium from forest soil samples heated at 65, 80, 95, 105, and 120 C, respectively, for 60 minutes. In all them, Clostridium was identified as the main H 2 -producing genus. ...
... It is generally known that several factors, e.g., pH and temperature (Kim et al., 2011), substrate concentration (Ning et al., 2013), source of inoculum and inoculum pretreatment method (Ravindran, Adav & Yang, 2010;Cai & Wang, 2016), and phase of microbial growth during the fermentation (i.e., lag, exponential or stationary phases) (Fang, Li & Zhang, 2006;Huang et al., 2010) can affect the microbial community, which in turn affect efficiency of hydrogen production process (O-Thong, 2017). Due to the differences in optimum conditions for hydrogen production observed for the acid-and acid-thermal pretreated biomass, it was interesting to investigate whether the microbial community in the two fermentations was different. ...
Background
Owing to the high growth rate, high protein and carbohydrate contents, and an ability to grow autotrophically, microalgal biomass is regarded as a promising feedstock for fermentative hydrogen production. However, the rigid cell wall of microalgae impedes efficient hydrolysis of the biomass, resulting in low availability of assimilable nutrients and, consequently, low hydrogen production. Therefore, pretreatment of the biomass is necessary in order to achieve higher hydrogen yield (HY). In the present study, acid-thermal pretreatment of Chlorella sp. biomass was investigated. Conditions for the pretreatment, as well as those for hydrogen production from the pretreated biomass, were optimized. Acid pretreatment was also conducted for comparison.
Results
Under optimum conditions (0.75% (v/v) H2SO4, 160 °C, 30 min, and 40 g-biomass/L), acid-thermal pretreatment yielded 151.8 mg-reducing-sugar/g-biomass. This was around 15 times that obtained from the acid pretreatment under optimum conditions (4% (v/v) H2SO4, 150 min, and 40 g-biomass/L). Fermentation of the acid-thermal pretreated biomass gave 1,079 mL-H2/L, with a HY of 54.0 mL-H2/g-volatile-solids (VS), while only 394 mL/L and 26.3 mL-H2/g-VS were obtained from the acid-pretreated biomass.
Conclusions
Acid-thermal pretreatment was effective in solubilizing the biomass of Chlorella sp. Heat exerted synergistic effect with acid to release nutrients from the biomass. Satisfactory HY obtained with the acid-thermal pretreated biomass demonstrates that this pretreatment method was effective, and that it should be implemented to achieve high HY.
... There are many different methods to treat mixed microflora inoculum. Pretreatment methods reported in most research works mainly include heat-shock [2,5,6] , acid [7,8], base [8], aeration, freezing and thawing, chloroform, sodium 2-bromoethanesulfonate or 2-bromoethanesulfonic acid [2], photocatalytic pretreatment [8,9] and ultrasonic treatments [8,10]. Heat shock is the most commonly used methods for treatment of mixed culture. ...
... Due to their high efficiency, Clostridium sp. are often used as pure culture to inoculate or bioaugment reactors. As they are spore-forming bacteria, their capacity to resist heat shock or other drastic pre-treatment enables to easily and efficiently select them from mixed inoculum, with long-term stability (Ravindran, Adav and Yang 2010;Park et al. 2014;Goud, Sarkar and Mohan 2014). Therefore, most of the research effort on H 2 production by dark fermentation has focused on Clostridia to date, and some relevant examples are provided below. ...
One of the most important biotechnological challenges is to develop environment friendly technologies to produce new sources of energy. Microbial production of biohydrogen through dark fermentation, by conversion of residual biomass, is an attractive solution for short-term development of bioH2 producing processes. Efficient biohydrogen production relies on complex mixed communities working in tight interaction. Species composition and functional traits are of crucial importance to maintain the ecosystem service. The analysis of microbial community revealed a wide phylogenetic diversity that contributes in different—and still mostly unclear—ways to hydrogen production. Bridging this gap of knowledge between microbial ecology features and ecosystem functionality is essential to optimize the bioprocess and develop strategies toward a maximization of the efficiency and stability of substrate conversion. The aim of this review is to provide a comprehensive overview of the most up-to-date biodata available and discuss the main microbial community features of biohydrogen engineered ecosystems, with a special emphasis on the crucial role of interactions and the relationships between species composition and ecosystem service. The elucidation of intricate relationships between community structure and ecosystem function would make possible to drive ecosystems toward an improved functionality on the basis of microbial ecology principles.
... There are many different methods to treat mixed microflora inoculum. Pretreatment methods reported in most research works mainly include heat-shock [2,5,6], acid [7,8], base [8], aeration, freezing and thawing, chloroform, sodium 2bromoethanesulfonate or 2-bromoethanesulfonic acid [2], photocatalytic pretreatment [8,9] and ultrasonic treatments [8,10]. Heat shock is the most commonly used methods for treatment of mixed culture. ...
In this study, batch biohydrogen production by co-digestion of raw rice straw and activated sewage sludge was investigated with different inoculum heat treatment, pH, S/X ratio (based on VS) and substrate sizes under mesophilic condition. In order to achieve a high bio-hydrogen yield and methanogens activity inhibition, heat treatment of inoculum was optimized at different exposure times (30, 45 & 60 min) and temperature ranges (80, 90 and 100 °C) prior to dark fermentation process. Collected data was analysed using response surface methodology (RSM). The heat treatment of inoculum at 100 °C for 60 min produced the highest bio-hydrogen yield of 14.22 NmL H2/g VS at concentration of 70.97% and Production of 0.073 NmL CH4/g VS at 0.17% concentration in total produced biogas. The raw rice straw was also co-digested with heat-treated inoculum at different ratios of volatile solids (2:1, 4:1 and 6:1) and initial pH (4, 4.75 and 5.5) as numerical variables and 4 categories of substrate size ((250–500 μm], (500 μm-2mm], (2–20 mm), [20–30 mm]). The highest bio-hydrogen yield of 14.70 NmL/g VS was recognized at the optimum initial pH of 5.01 and S/X ratio of 4.54:1 using 2–20 mm rice straw.
... Likewise, Wang et al. (2011) observed that increasing the temperature of heat pre-treatment from 80 C to 100 C resulted in decreased H 2 yields and concluded that overheating was not beneficial to H 2 production, an observation shared by Nissil€ a et al. (2011aNissil€ a et al. ( , 2011b). The two parameters of pre-treatment during HST are heating temperature varying from 40 to 120 C ( Wong et al., 2014b;Xiao and Liu, 2006;Baghchehsaraee et al., 2008;Guo et al., 2010b;Mu et al., 2007;Oh et al., 2003;Ravindran et al., 2010) and duration of heating ranging from 10 to 180 min ( Karadag et al., 2009;Hu and Chen, 2007;Nissil€ a et al., 2011a;Chairattanamanokorn et al., 2009;O-Thong et al., 2009;Sivaramakrishna et al., 2010;Assawamongkholsiri et al., 2013;Bakonyi et al., 2014;Vijayaraghavan et al., 2006) as summarised in Table 1. For more studies on the use of heat for inoculum pre-treatment, refer to the review done by Wong et al. (2014a). ...
Biohydrogen production from dark fermentation of lignocellulosic materials represents a huge potential in terms of renewable energy exploitation. However, the low hydrogen yield is currently hindering its development on industrial scale. This study reviewed various technologies that have been investigated for enhancing dark fermentative biohydrogen production. The pre-treatment technologies can be classified based on their applications as inoculum or substrates pre-treatment or they can be categorised into physical, chemical, physicochemical and biological based on the techniques used. From the different technologies reviewed, heat and acid pre-treatments are the most commonly studied technologies for both substrates and inoculum pre-treatment. Nevertheless, these two technologies need not necessarily be the most suitable since across different studies, a wide array of other emerging techniques as well as combined technologies have yielded positive findings. To date, there exists no perfect technology for either inoculum or substrate pre-treatment. Although the aim of inoculum pre-treatment is to suppress H2-consumers and enrich H2-producers, many sporulating H2-consumers survive the pre-treatment while some non-spore H2-producers are inhibited. Besides, several inoculum pre-treatment techniques are not effective in the long run and repeated pre-treatment may be required for continuous suppression of H2-consumers and sustained biohydrogen production. Furthermore, many technologies employed for substrates pre-treatment may yield inhibitory compounds that can eventually decrease biohydrogen production. Consequently, much research needs to be done to find out the best technology for both substrates and inoculum pre-treatment while also taking into consideration the energetic, economic and technical feasibility of implementing such a process on an industrial scale.
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... Bacilli are known to be good producers of biohydrogen where as Clostridia have iron dependent hydrogenase ([FeFe]-hydrogenase) as a dominant functional gene which has higher activity than [NiFe]-hydrogenase [25,29]. C. beijerinckii have the ability to yield higher H 2 by utilizing variety of carbon and nitrogen sources [30]. C. Acetobutyricum ferments carbohydrates to H 2 and carbon dioxide with acetate and butyrate as the main soluble metabolites [31]. ...
The effect of heat-shock treatment to selectively enrich acidogenic, H2 producing consortia was investigated for inoculum preparation and to control the process operation. Long term operation (520 days) in suspended-batch mode bioreactors illustrated relative efficiency and feasibility of heat-shock treated consortia (15.78 mol/kg CODR) in enhancing H2 production (3.31 mol/kg CODR) when compared to parent (control) consortia. On the contrary, substrate degradation was higher in the control operation (ξCOD, 62.86%; substrate degradation rate (SDR), 1.34 kg CODR/m3-day) compared to heat-shock operation (ξCOD, 52.6%; SDR, 1.10 kg CODR/m3-day). Heat-shock pretreatment has resulted in a marked fermentation pathway shift towards acetic-butyric acid type production. The microbial diversity illustrated dominance in the Clostridia class after applying heat-shock pretreatment. The redox catalytic currents and Tafel analysis strongly support the conclusion of an improved biocatalyst performance after pretreatment with regards to H2 production.
... In addition, the effects of pH and temperature on hydrogen production potential were studied with the most efficient H 2 producing enrichment. To the authors knowledge the effect of heat pretreatment on H 2 producing microbial communities has only been reported few times [23,24] and never for rumen fluid cultures. ...
Elevated temperatures (52, 60 and 65 °C) were used to enrich hydrogen producers on cellulose from cow rumen fluid. Methanogens were inhibited with two different heat treatments. Hydrogen production was considerable at 60 °C with the highest H2 yield of 0.44 mol-H2 mol-hexose−1 (1.93 mol-H2 mol-hexose-degraded−1) as obtained without heat treatment and with acetate and ethanol as the main fermentation products. H2 production rates and yields were controlled by cellulose degradation that was at the highest 21%. The optimum temperature and pH for H2 production of the rumen fluid enrichment culture were 62 °C and 7.3, respectively. The enrichments at 52 and 60 °C contained mainly bacteria from Clostridia family. At 52 °C, the bacterial diversity was larger and was not affected by heat treatments. Bacterial diversity at 60 °C remained similar between heat treatments, but decreased during enrichment. At 60 °C, the dominant microorganism was Clostridium stercorarium subsp. leptospartum.
For decades, reclamation of pesticide contaminated sites has been a challenging avenue. Due to increasing agricultural demand, the application of synthetic pesticides could not be controlled in its usage, and it has now adversely impacted the soil, water, and associated ecosystems posing adverse effects on human health. Agricultural soil and pesticide manufacturing sites, in particular, are one of the most contaminated due to direct exposure. Among various strategies for soil reclamation, ecofriendly microbial bioremediation suffers inherent challenges for large scale field application as interaction of microbes with the polluted soil varies greatly under climatic conditions. Methodically, starting from functional or genomic screening, enrichment isolation; functional pathway mapping, production of tensioactive metabolites for increasing the bioavailability and bio-accessibility, employing genetic engineering strategies for modifications in existing catabolic genes to enhance the degradation activity; each step-in degradation study has challenges and prospects which can be addressed for successful application. The present review critically examines the methodical challenges addressing the feasibility for restoring and reclaiming pesticide contaminated sites along with the ecotoxicological risk assessments. Overall, it highlights the need to fine-tune the available processes and employ interdisciplinary approaches to make microbe assisted bioremediation as the method of choice for reclamation of pesticide contaminated sites.
The study investigates the implications of waste feedstock, inoculum origin, and pretreatment on volatile fatty acids accumulation (VFA). The acidogenic fermentation of the feedstocks, rice mill effluent (RME), and brewery effluent (BE) was studied using untreated and pretreated (cyclic heat‐acid shock) brewery anaerobic sludge as inoculum. The pretreatment was successful in refining and stabilizing VFA production from the feedstocks. The fermentation of RME with pretreated sludge had an enhanced acetate yield of 0.37 ± 0.02 mgCOD/mgCOD, even to odd ratio of 20.97 ± 0.08 mg/mg and the highest butyrate yield of 0.39 ± 0.01 mgCOD/mgCOD compared to untreated system. The pretreated system had stability in COD and pH profile, while VFA content depends on the origin of inoculum. Pretreatment inhibited the carbon sinks and augmented acetate–butyrate type metabolism with stable performance. The fermentation of RME by pretreated sludge produced a higher even‐numbered VFAs and enhanced even to odd ratio in comparison with fermentation of BE, thereby affecting polymer composition and property.
Practitioner points
• The pretreated system had stable acidification, chemical oxygen demand, and pH profile.
• The pretreated system had higher acetate and butyrate yield compared to the untreated system.
• Rice mill effluent acidified with pretreated sludge had the highest even to odd ratio, 20.97 mg/mg.
• The even to odd ratio for acidification of brewery effluent was insignificant.
• Pretreatment, the origin of sludge, and the effluent had a regulatory effect on acidification.
The Palm Oil industry is growing faster, as the global demand for its products greatly overcomes its production. As palm oil production increases, also does its effluent, Palm Oil Mill Effluent (POME). POME is a complex effluent, which is not toxic, but due to its elevated organic content, it is considered extremely polluting. Generally, POME is submitted to both physical and chemical treatments before being discarded into receiving streams or other water bodies. This process might be costly for the industry and there 98is no income from effluent treatment, therefore another destination for this effluent is desirable. As an alternative, this residue could be exploited as raw material in biological processes, specially hydrogen and methane production, due to the presence of carbohydrates, lipids and proteins that might be metabolized during the dark fermentation process. Hydrogen can be used in different industries, such as chemical, biochemical and food industries. Methane is an important combustible, which could substitute natural gas and hydrogen as a very promising fuel. This review approaches sustainable and renewable processes for POME exploitation as raw material for renewable energy production.
Biohydrogen production using palm oil mill effluent (POME) as source of nutrients was evaluated via dark fermentation under mesophilic conditions. The effect of adding a plant enzyme preparation (PEP) extracted from dormant castor bean seeds (Ricinus communis L.) to improve the availability of nutrients from POME was also evaluated and carried out in two different approaches. The addition of PEP in a one step process, a simultaneous hydrolysis and fermentation, resulted in a significant reduction of adaptive phase by approximately 50%. Furthermore, hydrogen yield increased by 14% and hydrogen productivity by 48%. The two step process, which consists in a pre-hydrolysis followed by dark fermentation, promoted a hydrogen yield of 2.58 mmol H2/gDQO, reduced the adaptive phase by 75% and increased hydrogen productivity by 102% when compared to the original conditions of raw POME fermentation.
Simultaneous sulfide, nitrate and carbon removal in the effect of nitrate and/or sulfide addition using methanogenic cultures was investigated using batch cultures. The cultures supplemented with sulfide efficiently denitrified 50–500mgN l−1 nitrate to nitrogen gas. The mechanism proposed was denitrification occurred via autotrophic pathway first, followed by heterotrophic pathway. Nitrate reduction pathway slightly shifted to dissimilatory nitrate reduction to ammonia (DNRA) in cultures with no addition of sulfide, starting at intial nitrate concentration 250mgN l−1. Microbial community probing suggests that autotrophic denitrification were carried out by epsilon proteobacterium (Thiomicrospira denitrificans). The heterotrophic pathway was carried out by Thauera sp. in both cultures with and without the addition of sulfide.With the presence of nitrate, sulfide was oxidized to elemental sulfur in 32h after the assay began. Due to the activity of sulfate reducing strains, it was later recovered back to nearly initial concentration via heterotrophic oxidation pathway. Sole addition of nitrate at concentration 50–500mgN l−1 resulted in delay of methanogenesis. At concentration 750–1000mgN l−1, a complete suppression occurred. Addition of nitrate along with sulfide at concentration 100mg S l−1 decrease methane production, while at concentration 200mg S l−1, severe inhibition occurred. However, when sulfide was dose alone, the inhibition was not as severe as when it was added with nitrate. Inhibition was due to denitrification intermediates, mainly by accumulation of nitrite and nitrous oxide. Denitrification occurred first while at the same time inhibited methanogenesis.
Studies on how different functional strains interact in a microflora may include isolation of pure strains using conventional plating technique and then mix a few of the isolates before observing their growth in specific medium. As isolating pure strains that take part in the key function of industrial effluent purification via conventional method is impractical, convenient alternative approaches to screen essential microbial group that maintains desired function of a mixed population is desired. Such approaches can be employed to allow the selection and enrichment of so-called functional consortium with user-defined attributes for specific functions. This manuscript provides a review of various approaches to isolation and enrichment of microbial functional consortium in several biological processes. Consideration for the isolation and enrichment approaches and their applications are delineated. Challenges to the applications and further work are also outlined.
Cellulosic plant and waste materials are potential resources for fermentative hydrogen production. In this study, hydrogen producing, cellulolytic cultures were enriched from compost material at 52, 60 and 70°C. Highest cellulose degradation and highest H(2) yield were 57% and 1.4 mol-H(2) mol-hexose(-1) (2.4 mol-H(2) mol-hexose-degraded(-1)), respectively, obtained at 52°C with the heat-treated (80°C for 20 min) enrichment culture. Heat-treatments as well as the sequential enrichments decreased the diversity of microbial communities. The enrichments contained mainly bacteria from families Thermoanaerobacteriaceae and Clostridiaceae, from which a bacterium closely related to Thermoanaerobium thermosaccharolyticum was mainly responsible for hydrogen production and bacteria closely related to Clostridium cellulosi and Clostridium stercorarium were responsible for cellulose degradation.
Continuous, dark fermentative hydrogen production technology using mixed microflora at mesophilic temperatures may be suitable for commercial development. Clostridial-based cultures from natural sources have been widely used, but more information on the need for heat treatment of inocula and conditions leading to germination and sporulation are required. The amount of nutrients given in the literature vary widely. Hydrogen production is reported to proceed without methane production in the reactor in the pH range 4.5–6.7, with hydraulic retention times optimally between a few hours and 3 days depending on substrate. Higher substrate concentrations should be more energy-efficient but there are product inhibition limitations, for example from unionised butyric acid. Inhibition by H2 can be reduced by stirring, sparging or extraction through membranes. Of the reactor types investigated, while granules have the best performance with soluble substrate, for particulate feedstock biofilm reactors or continuous stirred tank reactors may be most successful. A second stage is required to utilise the fermentation end products which, when cost-effective reactors are developed, may be photofermentation or microbial fuel cell technologies. Anaerobic digestion is a currently-available technology and the two-stage process is reported to give greater conversion efficiency than anaerobic digestion alone.
We describe a new molecular approach to analyzing the genetic diversity of complex microbial populations. This technique is based on the separation of polymerase chain reaction-amplified fragments of genes coding for 16S rRNA, all the same length, by denaturing gradient gel electrophoresis (DGGE). DGGE analysis of different microbial communities demonstrated the presence of up to 10 distinguishable bands in the separation pattern, which were most likely derived from as many different species constituting these populations, and thereby generated a DGGE profile of the populations. We showed that it is possible to identify constituents which represent only 1% of the total population. With an oligonucleotide probe specific for the V3 region of 16S rRNA of sulfate-reducing bacteria, particular DNA fragments from some of the microbial populations could be identified by hybridization analysis. Analysis of the genomic DNA from a bacterial biofilm grown under aerobic conditions suggests that sulfate-reducing bacteria, despite their anaerobicity, were present in this environment. The results we obtained demonstrate that this technique will contribute to our understanding of the genetic diversity of uncharacterized microbial populations.
Production of hydrogen by anaerobes, facultative anaerobes, aerobes, methylotrophs, and photosynthetic bacteria is possible. Anaerobic Clostridia are potential producers and immobilized C. butyricum produces 2 mol H2/mol glucose at 50% efficiency. Spontaneous production of H2 from formate and glucose by immobilized Escherichia coli showed 100% and 60% efficiencies, respectively. Enterobactericiae produces H2 at similar efficiency from different monosaccharides during growth. Among methylotrophs, methanogenes, rumen bacteria, and thermophilic archae, Ruminococcus albus, is promising (2.37 mol/mol glucose). Immobilized aerobic Bacillus licheniformis optimally produces 0.7 mol H2/mol glucose. Photosynthetic Rhodospirillum rubrum produces 4, 7, and 6 mol of H2 from acetate, succinate, and malate, respectively. Excellent productivity (6.2 mol H2/mol glucose) by co-cultures of Cellulomonas with a hydrogenase uptake (Hup) mutant of R. capsulata on cellulose was found. Cyanobacteria, viz., Anabaena, Synechococcus, and Oscillatoria sp., have been studied for photoproduction of H2. Immobilized A. cylindrica produces H2 (20 ml/g dry wt/h) continually for 1 year. Increased H2 productivity was found for Hup mutant of A. variabilis. Synechococcus sp. has a high potential for H2 production in fermentors and outdoor cultures. Simultaneous productions of oxychemicals and H2 by Klebseilla sp. and by enzymatic methods were also attempted. The fate of H2 biotechnology is presumed to be dictated by the stock of fossil fuel and state of pollution in future.
Hydrogen gas (approximately 60% H(2)) was produced in a continuous flow bioreactor inoculated with heat-treated soil, and fed synthetic wastewater containing glucose (9.5 g l(-1)). The pH in the bioreactor was maintained at 5.5 to inhibit consumption of H(2) by methanogens. The objective of this study was to characterize bacterial communities in the reactor operated under two different hydraulic retention times (HRTs of 30-h and 10-h) and temperatures (30 degrees C and 37 degrees C). At 30-h HRT, the H(2) production rate was 80 ml h(-1) and yield was 0.91 mol H(2)/mol glucose. At 10-h HRT, the H(2) production rate was more than 5 times higher at 436 ml h(-1), and yield was 1.61 mol H(2)/mol glucose. Samples were removed from the reactor under steady-state conditions for PCR-based detection of bacterial populations by ribosomal intergenic spacer analysis (RISA). Populations detected at 30-h HRT were more diverse than at 10-h HRT and included representatives of Bacillaceae, Clostridiaceae, and Enterobacteriaceae. At 10-h HRT, only Clostridiaceae were detected. When the temperature of the 10-h HRT reactor was increased from 30 degrees C to 37 degrees C, the steady-state H(2) production rate increased slightly to 463 ml h(-1) and yield was 1.8 mol H(2)/mol glucose. Compared to 30 degrees C, RISA fingerprints at 37 degrees C from the 10-h HRT bioreactor exhibited a clear shift from populations related to Clostridium acidisoli (subcluster Ic) to populations related to Clostridium acetobutylicum (subcluster Ib).
The global temperature rose by 0.2 degrees C between the middle 1960's and 1980, yielding a warming of 0.4 degrees C in the past century. This temperature increase is consistent with the calculated greenhouse effect due to measured increases of atmospheric carbon dioxide. Variations of volcanic aerosols and possibly solar luminosity appear to be primary causes of observed fluctuations about the mean trend of increasing temperature. It is shown that the anthropogenic carbon dioxide warming should emerge from the noise level of natural climate variability by the end of the century, and there is a high probability of warming in the 1980's. Potential effects on climate in the 21st century include the creation of drought-prone regions in North America and central Asia as part of a shifting of climatic zones, erosion of the West Antarctic ice sheet with a consequent worldwide rise in sea level, and opening of the fabled Northwest Passage.
The rate of hydrogen production by the marine nonsulfur photosynthetic bacterium, Rhodovulum sp., increased with increasing light intensity. A light intensity of 1800 W/m² hydrogen production rate was achieved at the rate of 9.4 μmol/mg dry weight/h. The hydrogen production of this strain was enhanced by the addition of a small amount of oxygen (12 μmol O2/reactor). Intracellular ATP content was most efficiently accumulated under microaerobic, dark conditions. Hydrogen production rate by Rhodovulum sp. was investigated using a double-phase photobioreactor consisting of light and dark compartments. This rate was compared with data obtained using a conventional photobioreactor. Rhodovulum sp. produced hydrogen at a rate of 0.38 ± 0.03 μmol/mg dry weight/h under microaerobic conditions using the double-phase photobioreactor. The hydrogen production rate was four times greater under microaerobic conditions, as compared with anaerobic conditions using either type of photobioreactor. Hydrogen production using a double-phase photobioreactor was demonstrated continuously at the same rate for 150 h. © 2000 John Wiley & Sons, Inc. Biotechnol Bioeng 68: 647–651, 2000.
Cellulase production by submerged fermentation (SF) and solid state fermentation (SSF) is compared. In SF the average production level is 10 filter paper unit (FPU) /ml with a volumetric productivity of 100 FPU/1, h. In SSF, a production level of 100 FPU/gDW substrate with a volumetric productivity of 320 FPU/g, h may be reached. The production cost of cellulase in the crude fermentum by SF is about $ 20/kg cellulase, by SSF it is only $ 0.2/kg cellulase, if in situ fermentation is used. The enzyme in the SSF crude product is concentrated, thus it may be used directly in such agrobiotechnological applications as silage or feed additive, lignocellulose hydrolysis, and natural fiber (e.g. jute) processing.
Biological hydrogen production has been known for over a century and research directed at applying this process to a practical means of hydrogen fuel production has been carried out for over a quarter century. The various approaches that have been proposed and investigated are reviewed and critical limiting factors identified. The low energy content of solar irradiation dictates that photosynthetic processes operate at high conversion efficiencies and places severe restrictions on photobioreactor economics. Conversion efficiencies for direct biophotolysis are below 1% and indirect biophotolysis remains to be demonstrated. Dark fermentation of biomass or wastes presents an alternative route to biological hydrogen production that has been little studied. In this case the critical factor is the amount of hydrogen that can be produced per mole of substrate. Known pathways and experimental evidence indicates that at most 2–3mol of hydrogen can be obtained from substrates such as glucose. Process economics require that means be sought to increase these yields.
Trichoderma reesei LM-UC4, the parent strain, and its hypercellulolytic mutant LM-UC4E1 were co-cultured with Aspergillus niger ATCC 10864 in solid substrate fermentation on alkali-treated sugar cane for cellulolytic enzyme production. Bagasse was supplemented with either soymeal or with ammonium sulfate and urea, and fermented at 80% moisture content and 30°C. Mixed culturing produced better results with the inorganic supplement. The mutant strain was more responsive to mixed culturing than the parent strain when A. niger was the cooperating partner. In a mixed culture of the mutant with inorganic N-source, 10% more biomass, but 63% more cellulase, 85% more endoglucanase and 147% more β-glucosidase was produced than in single culture. Since co-culturing helped enzyme production more than growth, it appeared that synergistic interactions not directly related to growth were responsible for increased enzyme production. Although soymeal supplementation increased biomass production in the same mixed culture by 30%, it did not increase enzyme production. Mixed culturing is thus beneficial for the economic production of cellulases on nutritionally poor agricultural residues, without the need for supplementation with expensive organic supplements.
Hydrogen is the fuel of the future mainly due to its high conversion efficiency, recyclability and nonpolluting nature. Biological hydrogen production processes are found to be more environment friendly and less energy intensive as compared to thermochemical and electrochemical processes. They are mostly controlled by either photosynthetic or fermentative organisms. Till today, more emphasis has been given on the former processes. Nitrogenase and hydrogenase play very important role. Genetic manipulation of cyanobacteria (hydrogenase negative gene) improves the hydrogen generation. The paper presents a survey of biological hydrogen production processes. The microorganisms and biochemical pathways involved in hydrogen generation processes are presented in some detail. Several developmental works are discussed. Immobilized system is found suitable for the continuous hydrogen production. About 28% of energy can be recovered in the form of hydrogen using sucrose as substrate. Fermentative hydrogen production processes have some edge over the other biological processes.
The improvement of the yield of hydrogen at fermentative production using molasses as a substrate was investigated from two points of view; i.e. firstly, through the control of production pathway and secondly, by the condition of nutrient source for growing. It was shown that succinate, one of the products of Enterobacter aerogenes strain E.82005, was produced inductively by accumulation of CO2 which was produced simultaneously with acetate, ethanol and other products. The amount of residual NADH which is the base material of H2 evolution increased when Ar or H2 was blown into the culture liquid. From these results, it was concluded that the removal of CO2 from culture liquid effected the promotion of the yield of hydrogen. It was also shown that the existence of sufficient nitrogen source for bacterial growth was a necessary condition to get entire yield of hydrogen. The combination of experiments to remove CO2 and to give sufficient nitrogen source was found to yield 1.58 mol hydrogen from 1 mol sugar in terms of glucose.
Various medium components (carbon and nitrogen sources, iron, inoculum size) and environmental factors (initial pH and the agitation speed) were evaluated for their effects on the rate and the yield of hydrogen production by Clostridium saccharoperbutylacetonicum. Among the carbon sources assessed, cells grown on disaccharides (lactose, sucrose and maltose) produced on the average more than twice (2.81 mol-H2/mol sugar) as much hydrogen as monosaccharides (1.29 mol-H2/mol sugar), but there was no correlation between the carbon source and the production rate. The highest yield (2.83 mol/mol) was obtained in lactose and sucrose but the highest production rate (1.75 mmol/h) in sucrose. Using glucose as carbon source, yeast extract was the best nitrogen source. A parallel increase between the production rate and the yield was obtained by increasing glucose concentration up to 40 g/l (1.76 mol-H2/mol, 3.39 mmol/h), total nitrogen as yeast extract up to 0.1% (1.41 mol/mol, 1.91 mmol/h) and agitation up to 100 rev/min (1.66 mol-H2/mol, 1.86 mmol/h). On the other hand, higher production rates were favoured in preference to the yield at a neutral initial pH 7 (2.27 mmol/h), 1000 mg iron/l or more (1.99 mmol/h), and a larger inoculum size, 10%, (2.36 mmol/h) whereas an initial alkaline pH of 8.5 (1.72 mol/mol), a lower iron concentration of 25 mg/l (1.74 mol/mol) and smaller inoculum size, 1%, (1.85 mol/mol) promoted higher yield over
production rate.
A Clostridium butyricum strain, isolated from hydrogen-producing sewage sludge, was examined for its ability to produce H2 from sucrose-based medium under different medium composition, pH, and carbon substrate concentration. The strain, designated as C. butyricum CGS5, grew and produced hydrogen efficiently on iron-containing medium. Hydrogen started to evolve when cell growth entered mid-exponential phase and reached maximum production rate at the stationary phase. The optimal hydrogen production (5.3 l) and hydrogen yield (2.78 mol H2/mol sucrose) were obtained at an initial sucrose concentration of 20 g COD/l (17.8 g/l) and a pH of 5.5. However, the CGS5 strain attained its highest hydrogen production rate (209 ml/h/l) under a medium pH of 6.0. In comparison with pH 5.5, operation at pH 6.0 and 6.5 obtained higher cell growth rate and cell yield, but resulted in lower total hydrogen production and hydrogen yield. This is most likely due to rapid conversion of the carbon source into biomass, reducing the formation of hydrogen. Neither hydrogen production nor cell growth was detected when the strain was cultivated at pH 5.0. A sucrose concentration of 20 g COD/l gave the best hydrogen fermentation performance, whereas cell growth rate and hydrogen production rate both decreased when sucrose concentration was elevated to 30 g COD/l, suggesting that substrate inhibition may occur.
The merit of a hydrogen production system was investigated, where cells in the stationary phase of growth were treated as live enzymes, continually catalyzing hydrogen production in the absence of growth. Batch cultures of the purple photosynthetic bacteria Rhodospirillum rubrum UR2 were grown photoheterotrophically with succinate as the electron donor. Hydrogen evolved during growth, via the enzyme nitrogenase, at a rate of 21 mL gas L−1 culture h−1, and continued to evolve at high rates for about 70 h after cells had ceased growth. Hydrogen production stopped precisely when succinate was depleted from the medium. Upon replenishment of succinate to the cultures, hydrogen production resumed but cells did not grow further; however, the rate and yield of hydrogen production was lower with successive succinate additions than that measured during growth. These results suggest that hydrogen production is not strictly coupled to growth. Nevertheless, the results also establish the necessity for cell growth in order to maintain maximal hydrogen production rates. Supplementation of cultures with limited amounts of fresh growth medium, given in addition to the succinate replenishment, partially restored the hydrogen production rate and yield, along with a proportional increase in cell biomass. Results were confirmed in parallel experiments with Rhodopseudomonas palustris CGA009. A strategy is suggested for enhancing the biofuels to biomass production ratio under conditions of continuous cultivation with minimal cell growth (about 10% of the control), allowing a greater proportion of the cellular metabolic activity to be directed toward H2-production.
The effect of heat treatment at different temperatures on two types of inocula, activated sludge and anaerobically digested sludge, was investigated in batch cultures. Heat treatments were conducted at 65, 80 and 95 °C for 30 min. The untreated inocula produced less amount of hydrogen than the pretreated inocula, with lactic acid as the main metabolite. The maximum yields of 2.3 and 1.6 mol H2/mol glucose were achieved for the 65 °C pretreated anaerobically digested and activated sludges, respectively. Approximately a 15% decrease in yield was observed with increasing pretreatment temperature from 65 to 95 °C concomitant with an increase in butyrate/acetate ratio from 1.5 to 2.4 for anaerobically digested sludge. The increase of pretreatment temperature of activated sludge to 95 °C suppressed the hydrogen production by lactic acid fermentation. DNA analysis of the microbial community showed that the elevated pretreatment temperatures reduced the species diversity.
A hydrogen producing strain newly isolated from anaerobic sludge in an anaerobic bioreactor, was identified as Clostridium beijerinckii Fanp3 by 16S rDNA gene sequence analysis and detection by BioMerieux Vitek. The strain could utilize various carbon and nitrogen sources to produce hydrogen, which indicates that it has the potential of converting renewable wastes into hydrogen. In batch cultivations, the optimal initial pH of the culture medium was between 6.47 and 6.98. Using 0.15 M phosphate as buffer could alleviate the medium acidification and improve the overall performance of C. beijerinckii Fanp3 in hydrogen production. Culture temperature of 35 °C was established to be the most favorable for maximum rate of hydrogen production. The distribution of soluble metabolic products (SMP) was also greatly affected by temperature. Considering glucose as a substrate, the activation energy (Ea) for hydrogen production was calculated as 81.01 kcal/mol and 21.4% of substrate energy was recovered in the form of hydrogen. The maximal hydrogen yield and the hydrogen production rate were obtained as 2.52 mol/mol-glucose and 39.0 ml/g-glucose h−1, respectively. These results indicate that C. beijerinckii Fanp3 is an ideal candidate for the fermentative hydrogen production.
The effects of lactic acid bacteria (LAB) on hydrogen fermentation of organic waste were investigated. For this three hydrogen producing strains of Clostridium were cultured with two lactic acid bacteria, i.e. Lactobacillus paracasei and Enterococcus durans, which were isolated from the wastes generated in the bean curd manufacturing. The decrease or cessation of hydrogen production by Clostridium was caused by the addition of LAB. The supernatants of L. paracasei and E. durans suspensions also inhibited hydrogen production by Clostridium. This inhibition was partially destroyed in the presence of trypsin, which is a protease inactivating a bacteriocin. These results suggest that the inhibitory effect of lactic acid bacteria on hydrogen production was caused by bacteriocins excreted from LAB which have a deleterious effect on other bacteria. To suppress any effect by LAB, heat treatment of this waste was investigated as a possible pretreatment step. The inhibition of hydrogen production was reduced by heat treatment for at temperatures ranging from 50°C to 90°C. This means that a temperature of 50°C is already adequate to prevent growth of LAB.
We evaluated the influence of the operation temperature (mesophilic vs. thermophilic regime) of semicontinuous, acidogenic solid substrate anaerobic digestion (A-SSAD) of the organic fraction of municipal solid waste (OFMSW) at lab scale. The H2 percentage was higher in the thermophilic regime than in the mesophilic operation (58% and 42%, respectively). The H2 yield of thermophilic A-SSAD was significantly higher than in our mesophilic reactors (360 vs. 165 NmL H2/g VSrem) and other studies reported in the literature (range of 62–180 mL/g VS). Mesophilic A-SSAD showed a yield of 37% of the maximum yield based on 4 mol H2/mol hexose, while thermophilic A-SSAD exhibited a yield of 80% of the maximum yield. This result is similar to works with pure cultures of extremophile microorganisms where H2 yields of 83% of the maximum were reported. We found higher concentrations of acetic acid in the digestates of thermophilic A-SSAD, while butyrate was in higher proportion in mesophilic A-SSAD spent solids. The moderate-to-high yields obtained with the semicontinuous process used in this work are in disagreement with previous reports claiming that batch and semicontinuous processes are less efficient than continuous ones.
Anaerobic sewage sludge acclimated with sucrose was used as the seed in a batch experiment to investigate the carbon/nitrogen (C/N)-ratio effects on biological hydrogen production from sucrose. Experimental results indicated that the hydrogen production ability of the anaerobic microflora (dominated by Clostridium pasteurianum) in the sewage sludge was dependent on the influent C/N-ratio. At a C/N-ratio of 47, the hydrogen productivity and hydrogen production rate reached and , respectively. This increased by 500% and 80%, respectively, compared with the blank. Proper C/N-ratio on hydrogen production enhancement was accomplished by shifting the metabolic pathway. Strategies for optimal hydrogen production are also proposed.
Growth and hydrogen production by two extreme thermophiles during sugar fermentation was investigated. In cultures of Caldicellulosiruptor saccharolyticus grown on sucrose and Thermotoga elfii grown on glucose stoichiometries of of hydrogen and of acetate per mol C6-sugar unit were obtained. The hydrogen level was about 83% of the theoretical maximum. C. saccharolyticus and T. elfii reached maximum cell densities of 1.1×109 and 0.8×109 cells/ml, respectively, while their maximum hydrogen production rates were 11.7 and dry weight/h, respectively. For growth of C. saccharolyticus on sucrose, a biomass yield of sucrose and a YATP of 11.3–14.1 were calculated. Replacement of yeast extract by casamino acids, plus proline and vitamins in the medium of C. saccharolyticus resulted in similar yields of hydrogen production on sucrose, but diminished the rate by about 30%. Both yeast extract and tryptone were required by T. elfii, and appeared to function as sources of carbon, nitrogen and energy. In the absence of tryptone, T. elfii converted 26% of the glucose to another by-product, resulting in a lower yield of hydrogen. Growth of T. elfii ceased prior to glucose depletion, but the culture continued to ferment glucose to hydrogen and acetate until all glucose was consumed.
Hydrogen may be produced by a number of processes, including electrolysis of water, thermocatalytic reformation of hydrogen-rich organic compounds, and biological processes. Currently, hydrogen is produced, almost exclusively, by electrolysis of water or by steam reformation of methane. Biological production of hydrogen (Biohydrogen) technologies provide a wide range of approaches to generate hydrogen, including direct biophotolysis, indirect biophotolysis, photo-fermentations, and dark-fermentation. The practical application of these technologies to every day energy problems, however, is unclear. In this paper, hydrogen production rates of various biohydrogen systems are compared by first standardizing the units of hydrogen production and then by calculating the size of biohydrogen systems that would be required to power proton exchange membrane (PEM) fuel cells of various sizes.
A gram negative hydrogen producing facultative anaerobe was isolated and characterized as Enterobacter cloacae IIT-BT 08. Hydrogen yields by using this microorganism varied from substrate to substrate (2.2 mol/mol glucose, 6 mol/mol of sucrose and 5.4 mol/mol cellobiose considering 1% w/v substrate in MY medium). The maximum rate of hydrogen production achieved was at 36°C and initial pH 6.0. The maximum rate was 29.63 mmol/(g dry cell per h). The pH profiles of the fermentation broth under aerobic and anaerobic conditions were monitored and found to differ from each other particularly beyond the pH of 4.8. About 28% of substrate energy were recovered in the form of hydrogen using sucrose as a substrate.
Simple sugars, oligosaccharides, polysaccharides, and their derivatives, including the methyl ethers with free or potentially free reducing groups, give an orange-yellow color when treated with phenol and concentrated sulfuric acid. The reaction is sensitive and the color is stable. By use of this phenol-sulfuric acid reaction, a method has been developed to determine submicro amounts of sugars and related substances. In conjunction with paper partition chromatography the method is useful for the determination of the composition of polysaccharides and their methyl derivatives.
In this paper, experiments were conducted to investigate H2 production from glucose by mixed anaerobic cultures at various temperatures in the mesophilic range. Results showed that glucose degradation rate and efficiency, H2 yield, and growth rate of H2-producing bacteria all increased as the temperature increased from 33 to View the MathML source. However, the specific H2 production rate increased with increasing temperature from 33 to View the MathML source, then decreased as the temperature was further increased to View the MathML source. The distribution of aqueous products was also greatly influenced by temperature variation. H2 yield and growth rate of H2-producing cultures had a linear relationship with temperature. A modified Gompertz model was able to adequately describe the H2 production and microbial growth in the mesophilic range. The activation energies for H2 production and microbial growth were estimated as 107.66 and 204.77 kJ/mol, respectively.
A functional bacterial consortium that can effectively hydrolyze cellobiose and produce bio-hydrogen was isolated by a concentration-to-extinction approach. The sludge from a cattle feedlot manure composting plant was incubated with 2.5-20 g l(-1) cellobiose at 35 degrees C and pH 6.0. The microbial diversity of serially concentrated suspensions significantly decreased following increasing cellobiose concentration, finally leaving only two viable strains, Clostridium butyricum strain W4 and Enterococcus saccharolyticus strain. This consortium has a maximum specific hydrogen production rate of 2.19 mol H(2)molhexose(-1) at 5 g l(-1) cellobiose. The metabolic pathways shifted from ethanol-type to acetate-butyrate type as cellobiose concentration increased from 2.5 to >7 g l(-1). The concentration-to-extinction approach is effective for isolating functional consortium from natural microflora. In this case the functional strains of interest are more tolerant to the increased loadings of substrates than the non-functional strains.
The rate of hydrogen production by the marine nonsulfur photosynthetic bacterium, Rhodovulum sp., increased with increasing light intensity. A light intensity of 1800 W/m(2) hydrogen production rate was achieved at the rate of 9.4 micromol/mg dry weight/h. The hydrogen production of this strain was enhanced by the addition of a small amount of oxygen (12 micromol O(2)/reactor). Intracellular ATP content was most efficiently accumulated under microaerobic, dark conditions. Hydrogen production rate by Rhodovulum sp. was investigated using a double-phase photobioreactor consisting of light and dark compartments. This rate was compared with data obtained using a conventional photobioreactor. Rhodovulum sp. produced hydrogen at a rate of 0.38+/-0.03 micromol/mg dry weight/h under microaerobic conditions using the double-phase photobioreactor. The hydrogen production rate was four times greater under microaerobic conditions, as compared with anaerobic conditions using either type of photobioreactor. Hydrogen production using a double-phase photobioreactor was demonstrated continuously at the same rate for 150 h.
The effect of the iron concentration in the external environment on hydrogen production was studied using sucrose solution and the mixed microorganisms from a soybean-meal silo. The iron concentration ranged from 0 to 4,000 mg FeCl2 l(-1). The temperature was maintained at 37 degrees C. The maximum specific hydrogen production rate was found to be 24.0 ml g(-1) VSS h(-1) at 4,000 mg FeCl2 l(-1). The specific production rate of butyrate increased with increasing iron concentration from 0 to 20 mg FeCl2 l(-1) and decreased with increasing iron concentration from 20 to 4,000 mg FeCl2 l(-1). The maximum specific production rates of ethanol (682 mg g(-1) VSS h(-1)) and butanol (47.0 mg g(-1) VSS h(-1)) were obtained at iron concentrations of 5 and 3 mg FeCl2 l(-1), respectively. The maximum hydrogen production yield of 131.9 ml g(-1) sucrose was obtained at the iron concentration of 800 mg FeCl2 l(-1). The maximum yields of acetate (389.3 mg g(-1) sucrose), propionate (37.8 mg g(-1) sucrose), and butyrate (196.5 mg g(-1) sucros) were obtained at iron concentrations of 3, 200 and 200 mg FeCl2 l(-1), respectively. The sucrose degradation efficiencies were close to 1.0 when iron concentrations were between 200 and 800 mg FeCl2 l(-1). The maximum biomass production yield was 0.283 g VSS g(-1) sucrose at an iron concentration of 3,000 mg FeCl2 l(-1).
The effect of pH on the conversion of glucose to hydrogen by a mixed culture of fermentative bacteria was evaluated. At 36 degrees C, six hours hydraulic retention, over 90% of glucose was degraded at pH ranging 4.0-7.0, producing biogas and an effluent comprising mostly fatty acids. At the optimal pH of 5.5, the biogas comprised 64 +/- 2% of hydrogen with a yield of 2.1 +/- 0.1 mol-H2/mol-glucose and a specific production rate of 4.6 +/- 0.4 l-H2/(g-VSS day). The effluent was composed of acetate (15.3-34.1%) and butyrate (31.2-45.6%), plus smaller quantities of other volatile fatty acids and alcohols. The diversity of microbial communities increased with pH, based on 16S rDNA analysis by denaturing gradient gel electrophoresis (DGGE).
This study offers a novel and quick enrichment technology that can be used as a preliminary method to obtain a hydrogen-producing species from the biological sludge produced by wastewater treatment. The influences of acid-base enrichment (by sludge pH adjustment) on the anaerobic hydrogen-producing micro-organisms were investigated using serum bottle assays. The enrichment pH values were controlled at 3, 4, 5, 7, 10, 11 and 12 with 1 N hydrochloric acid and 1 N sodium hydroxide. For each enrichment pH, the cultivation pH values were controlled at 5, 6 and 7. Based on the experimental results, hydrogen accumulation from sludge with acid or base enrichment is higher than that of the control. The hydrogen-production potential of the sludge with acid or base enrichment is 200 and 333 times enhanced, compared with the control, when the enrichment pH is 10 and 3, respectively. The enhancement is due to a shortening of the micro-organisms' lag-time which occurs at a proper cultivation-pH level.
Virtually all members of the order Thermotogales have demonstrated the ability to produce hydrogen; however, some members of this order produce considerably greater quantities than others. With one representative of this order, Thermotoga neapolitana, we have consistently obtained accumulation of 25-30% hydrogen with 12-15% carbon dioxide as the only other prominent product in the batch reaction. In contradistinction to information widely disseminated in the literature, we have also found that most members of this order tolerate and appear to utilize the moderate amounts of oxygen present in the gaseous phase of batch reactors (6-12%), with no apparent decrease in hydrogen production. Hydrogen accumulation has been widely reported to inhibit growth of Thermotogales. While this may be true at very high hydrogen tensions, we have observed log phase bacterial morphology (rods) even in the presence of 25-35% hydrogen concentrations. To maximize hydrogen production and minimize production of hydrogen sulfide, inorganic sulfur donors are avoided and the cysteine concentration in the medium is increased. We and others have demonstrated that different members of the order Thermotogales utilize a wide variety of feedstocks, including complex carbohydrates and proteins. Thus, it appears that organisms within this order have the potential to utilize a variety of organic wastes and to cost-effectively generate hydrogen.
The biological production of hydrogen from the fermentation of different substrates was examined in batch tests using heat-shocked mixed cultures with two techniques: an intermittent pressure release method (Owen method) and a continuous gas release method using a bubble measurement device (respirometric method). Under otherwise identical conditions, the respirometric method resulted in the production of 43% more hydrogen gas from glucose than the Owen method. The lower conversion of glucose to hydrogen using the Owen protocol may have been produced by repression of hydrogenase activity from high partial pressures in the gastight bottles, but this could not be proven using a thermodynamic/rate inhibition analysis. In the respirometric method, total pressure in the headspace never exceeded ambient pressure, and hydrogen typically composed as much as 62% of the headspace gas. High conversion efficiencies were consistently obtained with heat-shocked soils taken at different times and those stored for up to a month. Hydrogen gas composition was consistently in the range of 60-64% for glucose-grown cultures during logarithmic growth but declined in stationary cultures. Overall, hydrogen conversion efficiencies for glucose cultures were 23% based on the assumption of a maximum of 4 mol of hydrogen/ mol of glucose. Hydrogen conversion efficiencies were similar for sucrose (23%) and somewhat lower for molasses (15%) but were much lower for lactate (0.50%) and cellulose (0.075%).
Sludge was granulated in a hydrogen-producing acidogenic reactor when operated at 26 degrees C, pH 5.5 treating a sucrose-rich wastewater. The influence of hydraulic retention time (HRT) and sucrose concentration on hydrogen production by the acidogenic granular sludge was investigated at a constant loading rate of 25 g-sucrose/(l x day). Results show that the gas composition was not greatly influenced by HRT or sucrose concentration. The hydrogen accounted for 57% to 68% of the biogas at HRT ranging 4.6-28.6 h and sucrose concentration ranging 4,800-29,800 mg/l. However, the hydrogen yield was more dependent on HRT and sucrose concentration. It ranged from 0.19 to 0.27 l/g-sucrose with the maximum yield occurring at HRT 13.7 h and sucrose concentration 14,300 mg/l in the wastewater. The acidified effluent was composed of volatile fatty acids and alcohols. The predominant products were butyrate (59-68%) and acetate (10-25%), plus smaller amounts of i-butyrate, valerate, i-valerate, caproate, methanol, ethanol, propanol, and butanol. The sludge yield averaged 0.2 g-VSS/g-sucrose. The carbon balance was 98-107% throughout the study.
Hydrogen gas can be recovered from the microbial fermentation of organic substrates at high concentrations when interspecies hydrogen transfer to methanogens is prevented. Two techniques that have been used to limit methanogenesis in mixed cultures are heat treatment, to remove nonsporeforming methanogens from an inoculum, and low pH during culture growth. We found that high hydrogen gas concentrations (57-72%) were produced in all tests and that heat treatment (HT) of the inoculum (pH 6.2 or 7.5) produced greater hydrogen yields than low pH (6.2) conditions with a nonheat-treated inoculum (NHT). Conversion efficiencies of glucose to hydrogen (based on a theoretical yield of 4 mol-H2/mol-glucose) were as follows: 24.2% (HT, pH = 6.2), 18.5% (HT, pH = 7.5), 14.9% (NHT, pH = 6.2), and 12.1% (NHT, pH = 7.5). The main products of glucose (3 g-COD/L) utilization (> or = 99%) in batch tests were acetate (3.4-24.1%), butyrate (6.4-29.4%), propionate (0.3-12.8%), ethanol (15.4-28.8%), and hydrogen (4.0-8.1%), with lesser amounts of acetone, propanol, and butanol (COD basis). Hydrogen gas phase concentrations in all batch cultures reached a maximum of 57-72% after 30 h but thereafter rapidly declined to nondetectable levels within 80 h. Separate experiments showed substantial hydrogen losses could occur via acetogenesis and that heat treatment did not prevent acetogenesis. Heat treatment consistently eliminated the production of measurable concentrations of methane. The disappearance of ethanol produced during hydrogen production was likely due to acetic acid production as thermodynamic calculations show that this reaction is spontaneous once hydrogen is depleted. Overall, these results show that low pH was, without heat treatment, sufficient to control hydrogen losses to methanogens in mixed batch cultures and suggest that methods will need to be found to limit acetogenesis in order to increase hydrogen gas yields by batch cultures.
Biological production of H(2) has received considerable attention lately. The present study was undertaken to observe the effects of substrate/seeding ratios (S(0)/X(0)) on batch H(2) generation. The H(2)-producing seeding spores were obtained from the heat treatment (88 degrees C for 12h) of the compost from a grass composting facility. A dehydrated brewery mixture was used as feed substrate. The results indicate that the pattern of the cumulative H(2) production with time is similar to the growth curve with a typical lag, exponential and stationary phase; the results were successfully modeled with a modified Gompertz equation. It appears that maximum H(2) yield potential (27ml g(-1)COD(added)) occurs at an S(0)/X(0) ratio of about 4, whereas the maximum specific H(2) yield (205ml g(-1) VSSd(-1)) occurs at approximately S(0)/X(0)=3. The S(0)/X(0) ratios higher than 4 would inhibit H(2) production. An attempt was made to waste a certain amount of reactor content and replaced it with fresh substrate in order to enhance H(2) production. After this medium replacement, the H(2) production was initially inhibited and the system then exhibited a long lag before it reached an active H(2) production stage. For a continuous-stirred tank-reactor (CSTR) system, the results of replacing 25% of the reactor content indicate that there is still a lag time before a sudden increase in H(2) production after the addition of the new substrate feed. The major low molecular weight acids identified are HAc and HBu with total volatile acids of about 6000-8000mg l(-1). The ratio of HAc/HBu in the present study is relatively constant (about 5) and appears not significantly affected by the medium replacement. The concentration of total alcohols is about 2000mg l(-1). All in all, the CSTR system is able to recover to its previous performance after such a dramatic 25% medium replacement.
The effect of conditioning for a variety of inoculums on fermentative hydrogen production was investigated. In addition, the effects of pH condition on hydrogen fermentation and bacterial community were investigated. The effect of conditioning on hydrogen production was different depending on the inoculum types. An appreciable hydrogen production was shown with anaerobic digested sludge and lake sediment without conditioning, however, no hydrogen was produced when refuse compost and kiwi grove soil were used as inoculums without conditioning. The highest hydrogen production was obtained with heat-conditioned anaerobic digested sludge, almost the same production was also obtained with unconditioned digested sludge. The pH condition considerably affected hydrogen fermentation, hydrogen gas was efficiently produced with unconditioned anaerobic sludge when the pH was controlled at 6.0 throughout the culture period and not when only the initial pH was adjusted to 6.0 and 7.0. Hydrogen production decreased when the culture pH was only adjusted at the beginning of each batch in continuous batch culture, and additionally, bacterial community varied with the change in hydrogen production. It was suggested that Clostridium and Coprothermobacter species played important role in hydrogen fermentation, and Lactobacillus species had an adverse effect on hydrogen production.
A mesophilic unsaturated flow (trickle bed) reactor was designed and tested for H2 production via fermentation of glucose. The reactor consisted of a column packed with glass beads and inoculated with a pure culture (Clostridium acetobutylicum ATCC 824). A defined medium containing glucose was fed at a flow rate of 1.6 mL/min (0.096 L/h) into the capped reactor, producing a hydraulic retention time of 2.1 min. Gas-phase H2 concentrations were constant, averaging 74 +/- 3% for all conditions tested. H2 production rates increased from 89 to 220 mL/hL of reactor when influent glucose concentrations were varied from 1.0 to 10.5 g/L. Specific H2 production rate ranged from 680 to 1270 mL/g glucose per liter of reactor (total volume). The H2 yield was 15-27%, based on a theoretical limit by fermentation of 4 moles of H2 from 1 mole of glucose. The major fermentation by-products in the liquid effluent were acetate and butyrate. The reactor rapidly (within 60-72 h) became clogged with biomass, requiring manual cleaning of the system. In order to make long-term operation of the reactor feasible, biofilm accumulation in the reactor will need to be controlled through some process such as backwashing. These tests using an unsaturated flow reactor demonstrate the feasibility of the process to produce high H2 gas concentrations in a trickle-bed type of reactor. A likely application of this reactor technology could be H2 gas recovery from pre-treatment of high carbohydrate-containing wastewaters.
Bacillus coagulans strain IIT-BT S1 isolated from anaerobically digested activated sewage sludge was investigated for its ability to produce H(2) from glucose-based medium under the influence of different environmental parameters. At mid-exponential phase of cell growth, H(2) production initiated and reached maximum production rate in the stationary phase. The maximal H(2) yield (2.28 mol H(2)/molglucose) was recorded at an initial glucose concentration of 2% (w/v), pH 6.5, temperature 37 degrees C, inoculum volume of 10% (v/v) and inoculum age of 14 h. Cell growth rate and rate of hydrogen production decreased when glucose concentration was elevated above 2% w/v, indicating substrate inhibition. The ability of the organism to utilize various carbon sources for H(2) fermentation was also determined.
Aerobic granules effectively degrade phenol at high concentrations. This work cultivated aerobic granules that can degrade phenol at a constant rate of 49 mg-phenol/g x VSS/h up to 1,000 mg/L of phenol. Fluorescent staining and confocal laser scanning microscopy (CLSM) tests demonstrated that an active biomass was accumulated at the granule outer layer. A strain with maximum ability to degrade phenol and a high tolerance to phenol toxicity isolated from the granules was identified as Candida tropicalis via 18S rRNA sequencing. This strain degrades phenol at a maximum rate of 390 mg-phenol/g x VSS/h at pH 6 and 30 degrees C, whereas inhibitory effects existed at concentrations >1,000 mg/L. The Haldane kinetic model elucidates the growth and phenol biodegradation kinetics of the C. tropicalis. The fluorescence in situ hybridization (FISH) and CLSM test suggested that the Candida strain was primarily distributed throughout the surface layer of granule; hence, achieving a near constant reaction rate over a wide range of phenol concentration. The mass transfer barrier provided by granule matrix did not determine the reaction rates for the present phenol-degrading granule.
Thirty five bacterial isolates from diverse environmental sources such as contaminated food, nitrogen rich soil, activated sludges from pesticide and oil refineries effluent treatment plants were found to belong to Bacillus, Bordetella, Enterobacter, Proteus, and Pseudomonas sp. on the basis of 16S rRNA gene sequence analysis. Under dark fermentative conditions, maximum hydrogen (H(2)) yields (mol/mol of glucose added) were recorded to be 0.68 with Enterobacter aerogenes EGU16 followed by 0.63 with Bacillus cereus EGU43 and Bacillus thuringiensis EGU45. H(2) constituted 63-69% of the total biogas evolved. Out of these 35 microbes, 18 isolates had the ability to produce polyhydroxybutyrate (PHB), which varied up to 500 mg/l of medium, equivalent to a yield of 66.6%. The highest PHB yield was recorded with B. cereus strain EGU3. Nine strains had high hydrolytic activities (zone of hydrolysis): lipase (34-38 mm) -Bacillus sphaericus strains EGU385, EGU399 and EGU542; protease (56-62 mm) -Bacillus sp. strains EGU444, EGU447 and EGU445; amylase (23 mm) -B. thuringiensis EGU378, marine bacterium strain EGU409 and Pseudomonas sp. strain EGU448. These strains with high hydrolytic activities had relatively low H(2) producing abilities in the range of 0.26-0.42 mol/mol of glucose added and only B. thuringiensis strain EGU378 had the ability to produce PHB. This is the first report among the non-photosynthetic microbes, where the same organism(s) -B. cereus strain EGU43 and B. thuringiensis strain EGU45, have been shown to produce H(2) - 0.63 mol/mol of glucose added and PHB - 420-435 mg/l medium.
In this study, cellulose hydrolysis activity of two mixed bacterial consortia (NS and QS) was investigated. Combination of NS culture and BHM medium exhibited better hydrolytic activity under the optimal condition of 35 degrees C, initial pH 7.0, and 100rpm agitation. The NS culture could hydrolyze carboxymethyl cellulose (CMC), rice husk, bagasse and filter paper, among which CMC gave the best hydrolysis performance. The CMC hydrolysis efficiency increased with increasing CMC concentration from 5 to 50g/l. With a CMC concentration of 10g/l, the total reducing sugar (RS) production and the RS producing rate reached 5531.0mg/l and 92.9mg/l/h, respectively. Furthermore, seven H2-producing bacterial isolates (mainly Clostridium species) were used to convert the cellulose hydrolysate into H2 energy. With an initial RS concentration of 0.8g/l, the H2 production and yield was approximately 23.8ml/l and 1.21mmol H2/g RS (0.097mmol H2/g cellulose), respectively.
Standard methods for the examination of water and wastewater
- Apha
APHA. Standard methods for the examination of water and wastewater. 20th ed. Washington DC, USA: American Public Health Association; 1998.
Microaerobic hydrogen production by photosynthetic bacteria in a double-phase photobioreactor
- Matsunaga
Conversion of chitinous wastes to hydrogen gas by Clostridium paraputrificum M-21
- Evvyernie
Biological hydrogen production measured in batch anaerobic respirometers
- Logan