Anthrahydroquinone-2,6-disulfonate increases the rate of hydrogen production during Clostridium beijerinckii fermentation with glucose,xylose, and cellobiose

Fermentative hydrogen production is considered a reasonable alternative for generating H₂ as an energy carrier for electricity production using hydrogen fuel cells. The kinetics of hydrogen production from glucose, xylose and cellobiose were investigated using pure culture Clostridium beijerinckii NCIMB 8052. Adding anthrahydroquinone-2,6-disulfonate (AH₂QDS) at concentrations ranging from 100 μM to 500 μM increased the hydrogen production rates from 0.80 to 1.35 mmol/L-hr to 1.20–2.70 mmol/L-hr with glucose, xylose, or cellobiose as the primary substrates. AH₂QDS amendment also increased the substrate utilization rate and biomass growth rate by at least two times. These findings suggest that adding hydroquinone reducing equivalents influence cellular metabolism with hydrogen production rate, substrate utilization rate, and growth rate being simultaneously affected. Resting cell suspensions were conducted to investigate the influence of AH₂QDS on the hydrogen production rate from glyceraldehyde 3-phosphate, which is a shared intermediate in both glycolysis and pentose phosphate pathway. Data demonstrated that hydrogen production rate increased by 1.5 times when glyceraldehyde 3-phosphate was the sole carbon source, suggesting that the hydroquinone may alter reactions starting with or after glyceraldehyde 3-phosphate in central metabolism. These data demonstrate that adding hydroquinones increased overall metabolic activity of C. beijerinckii. This will eventually increase the efficiency of industrial scale production once appropriate hydroquinone equivalents are identified that work well in large-scale operations, since fermentation rate is one of the two critical factors (production rate and yield) influencing efficiency and cost.     

Characterization on hydrogen production performance of a newly isolated Clostridium beijerinckii YA001 using xylose

A mesophilic bacterium was isolated from cow manure, and identified as Clostridium beijerinckii YA001 based on 16S rRNA gene sequence calculation using MEGA 5.0 and biochemical tests. This strain showed the ability to digest a wide range of carbon and nitrogen sources for hydrogen production, and it showed high hydrogen production performance using xylose as substrate. The optimum parameters for bio-hydrogen production in batch tests were pH 8.0, 1% substrate concentration, 40 °C and yeast extract as nitrogen source. The maximum hydrogen yield and the hydrogen production rate were obtained at 2.31 mol/mol xylose and 311.3 mL H₂/(Lh), respectively. These results indicate that C. beijerinckii YA001 is an ideal candidate for fermentative hydrogen production.     

Isolation and characterization of high hydrogen-producing strain Clostridium beijerinckii PS-3 from fermented oil palm sap

Felled oil palm trunk (OPT) (25 years old) is an abundant biomass in Southern Thailand. The OPT composition was 31.28–42.85% cellulose, 19.73–25.56% hemicellulose, 10.74–18.47% lignin, 1.63–2.25% protein, 1.60–1.83% fat, 1.12–1.35% ash and trace amount of minerals (0.01–0.40%). Oil palm sap extracted from OPT was found to contain 15.72 g/L glucose, 2.25 g/L xylose, and 0.086 g/L arabinose. A total of twenty samples from hot springs (45–75 °C and pH 6.5–8.4), oil palm sap and palm oil mill effluent were enriched for isolation of hydrogen-producing bacteria. The highest hydrogen-producing strain was isolated from oil palm sap and identified as Clostridium beijerinckii PS-3 using biochemical test and 16S rRNA gene analysis. Among various carbon sources tested, glucose, xylose, starch and cellulose were the preferred substrates for hydrogen production. The strain PS-3 could produce the maximum hydrogen yield of 140.9 ml H₂/g total sugar and the cumulative hydrogen production of 1973  ml/L-oil palm sap. Therefore, C. beijerinckii PS-3 is a potential candidate for fermentative hydrogen production from mixed sugars of the oil palm sap.     

Biological hydrogen production by extremely thermophilic novel bacterium Thermoanaerobacter mathranii A3N isolated from oil producing well

Hydrogen producing novel bacterial strain was isolated from formation water from oil producing well. It was identified as Thermoanaerobacter mathranii A3N by 16S rRNA gene sequencing. Hydrogen production by novel strain was pH and substrate dependent and favored pH 8.0 for starch, pH 7.5 for xylose and sucrose, pH 8.0–9.0 for glucose fermentation at 70 °C. The highest H₂ yield was 2.64 ± 0.40 mol H₂ mol glucose at 10 g/L, 5.36 ± 0.41 mol H₂ mol – sucrose at 10 g/L, 17.91 ± 0.16 mmol H₂ g – starch at 5 g/L and 2.09 ± 0.21 mol H₂ mol xylose at 5 g/L. The maximum specific hydrogen production rates 6.29 (starch), 9.34 (sucrose), 5.76 (xylose) and 4.89 (glucose) mmol/g cell/h. Acetate-type fermentation pathway (approximately 97%) was found to be dominant in strain A3N, whereas butyrate formation was found in sucrose and xylose fermentation. Lactate production increased with high xylose concentrations above 10 g/L.     

Evaluation of continuous biohydrogen production from enzymatically treated cornstalk hydrolysate

Enzymatically treated cornstalk hydrolysate was tested as substrate for H₂ production by Thermoanaerobacterium thermosaccharolyticum W16 in a continuous stirred tank reactor. The performance of strain W16 to ferment the main components of hydrolysate, mixture of glucose and xylose, in continuous culture was conducted at first, and then T. thermosaccharolyticum W16 was evaluated to ferment fully enzymatically hydrolysed cornstalk to produce H₂ in continuous operation mode. At the dilution rate of 0.020 h⁻¹, the H₂ yield and production rate reached a maximum of 1.9 mol H₂ mol⁻¹ sugars and 8.4 mmol H₂ L⁻¹ h⁻¹, respectively, accompanied with the maximum glucose and xylose utilizations of 86.3% and 77.6%. Continuous H₂ production from enzymatically treated cornstalk hydrolysate in this research provides a new direction for economic, efficient, and harmless H₂ production.     

Enhanced hydrogen production from xylose and bamboo stalk hydrolysate by overexpression of xylulokinase and xylose isomerase in Klebsiella oxytoca HP1

Biofuels production from lignocellulose hydrolysates by microbe fermentation has merited attention because of the mild reaction conditions involved and the clean nature of the process. In this work, xylulokinase (XK) and xylose isomerase (XI) were overexpressed in Klebsiella oxytoca HP1 to enhance hydrogen production by the fermentation of xylose. The recombinant strains exhibited higher enzyme activity of XI or XK compared with the wild strain. Hydrogen production from pure xylose, xylose/glucose mixtures and bamboo stalk hydrolysate was significantly enhanced with the overexpression of XI and XK in K. oxytoca HP1 in terms of total hydrogen yield (THY), hydrogen yield per mole substrate (HYPM) and hydrogen production rate (HPR). The HYPM of K. oxytoca HP1/xylB and K. oxytoca HP1/xylA reached 1.93 ± 0.05 and 2.46 ± 0.05 mol H₂/mol xylose, respectively in pure xylose, while the value for the wild strain was 1.68 ± 0.04 mol H₂/mol xylose. The xylose consumption rate (XCR) for the recombinant strain was significantly higher than that for the wild strain, particularly in the early stage of fermentation. Relative to the wild type, hydrogen yield (HY) from 1 g of preprocessed bamboo powder of HP1/xylB and HP1/xylA increased by 33.04 and 41.31%, respectively. It was concluded that overexpression of XK or XI was able to promote hydrogen production from xylose and xylose/glucose mixtures by simultaneously increasing the utilization efficiency of xylose and weakening the inhibitory effect of glucose on xylose use. In addition, the results indicated that overexpression technology was an effective way to further increase hydrogen production from lignocellulosic hydrolysates.     

Statistical optimization of fermentative hydrogen production from xylose by newly isolated Enterobacter sp. CN1

Statistical experimental designs were applied for the optimization of medium constituents for hydrogen production from xylose by newly isolated Enterobacter sp. CN1. Using Plackett–Burman design, xylose, FeSO₄ and peptone were identified as significant variables which highly influenced hydrogen production. The path of steepest ascent was undertaken to approach the optimal region of the three significant factors. These variables were subsequently optimized using Box–Behnken design of response surface methodology (RSM). The optimum conditions were found to be xylose 16.15 g/L, FeSO₄ 250.17 mg/L, peptone 2.54 g/L. Hydrogen production at these optimum conditions was 1149.9 ± 65 ml H₂/L medium. Under different carbon sources condition, the cumulative hydrogen volume were 1217 ml H₂/L xylose medium, 1102 ml H₂/L glucose medium and 977 ml H₂/L sucrose medium; the maximum hydrogen yield were 2.0 ± 0.05 mol H₂/mol xylose, 0.64 mol H₂/mol glucose. Fermentative hydrogen production from xylose by Enterobacter sp. CN1 was superior to glucose and sucrose.     

Impact of regulated pH on proto scale hydrogen production from xylose by an alkaline tolerant novel bacterial strain, Enterobacter cloacae DT-1

A hydrogen producing facultative anaerobic alkaline tolerant novel bacterial strain was isolated from crude oil contaminated soil and identified as Enterobacter cloacae DT-1 based on 16S rRNA gene sequence analysis. DT-1 strain could utilize various carbon sources; glycerol, CMCellulose, glucose and xylose, which demonstrates that DT-1 has potential for hydrogen generation from renewable wastes. Batch fermentative studies were carried out for optimization of pH and Fe²⁺ concentration. DT-1 could generate hydrogen at wide range of pH (5–10) at 37 °C. Optimum pH was; 8, at which maximum hydrogen was obtained from glucose (32 mmol/L), when used as substrate in BSH medium containing 5 mg/L Fe²⁺ ion. Decrease in hydrogen partial pressure by lowering the total pressure in the fermenter head space, enhanced the hydrogen production performance of DT-1 from 32 mmol H₂/L to 42 mmol H₂/L from glucose and from 19 mmol H₂/L to 33 mmol H₂/L from xylose. Hydrogen yield efficiency (HY) of DT-1 from glucose and xylose was 1.4 mol H₂/mol glucose and 2.2 mol H₂/mol xylose, respectively. Scale up of batch fermentative hydrogen production in proto scale (20 L working volume) at regulated pH, enhanced the HY efficiency of DT-1 from 2.2 to 2.8 mol H₂/mol xylose (1.27 fold increase in HY from laboratory scale). 84% of maximum theoretical possible HY efficiency from xylose was achieved by DT-1. Acetate and ethanol were the major metabolites generated during hydrogen production.     

Comparative study on biohydrogen production by newly isolated Clostridium butyricum SP4 and Clostridium beijerinckii SP6

The hydrogen-producing bacteria SP4 and SP6 were isolated from the compost and identified by 16S rRNA gene sequencing as Clostridium butyricum and Clostridium beijerinckii, respectively. A comparative study on the biohydrogen-producing activity of the isolated strains was carried out using mono-, di- and tri-saccharides belonging to both hexoses (maltose, glucose, mannose, fructose, lactose, galactose, sucrose, raffinose, cellobiose) and pentoses (xylose). To assess the biotechnological significance, real wastewater rich in sugars (cheese whey, confectionery wastewater, sugar beet processing wastewater) was also used as a substrate. C. butyricum SP4 fermented sugars with a yield of 0.93–1.52 mol H₂/mol hexose (pentose); the maximum yield was obtained from fructose, the minimum – from raffinose and cellobiose. The most preferred substrate for C. beijerinckii SP6 was sucrose with a yield of 1.76 mol H₂/mol hexose, while cellobiose yielded only 0.64 mol H₂/mol hexose. Overall, the efficiency of converting wastewater to H₂ by C. butyricum SP4 was also slightly lower (66–93 ml H₂/g chemical oxygen demand (COD)) than that of C. beijerinckii SP6 (76–103 ml H₂/g COD). Even though the main soluble metabolite products (SMPs) for both isolates were acetate and butyrate, C. butyricum SP4 also produced a significant amount of ethanol (up to 21.5% of SMPs) and formate (up to 32.5% of SMPs), and C. beijerinckii SP6 – lactate (up to 25% of SMPs). A distinctive feature of C. beijerinckii SP6 was a significantly lower (almost 2 times) yield of SMPs, while C. butyricum SP4 had a higher rate of H₂ production according to the results obtained from the kinetic study using the modified Gompertz equation and the first order equation. Analysis of Spearman's rank correlation coefficients revealed a statistically significant relationship between the kinetic parameters of H₂ production and the concentration of butyrate and the final pH of the medium for C. butyricum SP4, and with the concentration of ethanol for C. beijerinckii SP6. These findings provide valuable information on the metabolic capabilities of the most studied hydrogen-producing representatives of the Clostridium genus for their use in optimizing the technology for biohydrogen production by dark fermentation of various organic wastes.     

Hydrogen and ethanol fermentation of various carbon sources by immobilized Escherichia coli (XL1-Blue)

In this study, hydrogen and ethanol production by a facultative anaerobic bacterium Escherichia coli XL1-Blue immobilized in calcium-alginate beads have been investigated. Batch fermentations were carried out at mesophilic temperature (35 °C) and an initial cultivation pH of 6.5. Firstly, the influence of biomass concentration in terms of dry cell weight (expressed in g DCW/L, range 0.2–1.0) was investigated using fructose (5 g/L) as a carbon source. The peak hydrogen yield (HY) of 1.17 mol-H₂/mol-fructose_utilized was obtained at an initial cell concentration of 0.4 g DCW/L. The hydrogen production potential of other simple carbon sources (glucose and xylose) was evaluated at this optimized cell concentration and peak HY values were attained as 0.96 mol-H₂/mol-glucose_utilized and 0.69 mol-H₂/mol-xylose_utilized, respectively. In addition, utilization of the beverage wastewater (BWW) showed the peak cumulative hydrogen production and ethanol concentration of 120 mL and 5.65 g/L, attained at the substrate concentration of 20 g_{(glucose equivalent)}/L. However, peak HY (1.65 mol-H₂/mol-_{glucose eqivalent utilized}) was observed at low substrate concentration of 5 g_{(glucose equivalent)}/L. The percentage of sugar utilization of BWW was ranged between 80 and 96.     

Biohydrogen production from wastewater by Clostridium beijerinckii: Effect of pH and substrate concentration

An investigation of biological hydrogen production from glucose by Clostridium beijerinckii was conducted in a synthetic wastewater solution. A study examining the effect of initial pH (range 5.7–6.5) and substrate loading (range 1–3 g COD/L) on the specific conversion and hydrogen production rate has shown interaction behaviour between the two independent variables. Highest conversion of 10.3 mL H₂/(g COD/L) was achieved at pH of 6.1 and glucose concentration of 3 g COD/L, whereas the highest production rate of 71 mL H₂/(h L) was measured at pH 6.3 and substrate loading of 2.5 g COD/L. In general, there appears to be a strong trend of increasing hydrogen production rate with an increase in both substrate concentration and pH. Butyrate (14–63%), formate (10–45%) and ethanol (16–40%) were the main soluble products with other volatile fatty acids and alcohols present in smaller quantities.     

Potential for using enriched cultures and thermotolerant bacterial isolates for production of biohydrogen from oil palm sap and microbial community analysis

A limited number of bacteria can convert oil palm (Elaeis guineensis) sap to hydrogen with satisfactory yield and productivity. In this study, a total of 18 fermentative enriched cultures and 36 newly isolated thermotolerant bacterial strains were compared for hydrogen production from oil palm (OP) sap. The new isolates were obtained from hot springs, palm oil mill effluent and oil palm sap. The test was conducted in three steps: (i) a test for hydrogen production from mixed substrates (cellulose, starch, xylose, and glucose) and OP sap; (ii) a test for substrate concentration tolerance; and (iii) a test for thermotolerance. Five enriched candidates for each of the hydrogen producers were selected according to the criteria defined for the screening test. The hydrogen production of these selected bacterial strains from hot springs were cultivated in batch fermentation of oil palm sap at room temperature (30 ± 2 °C). Five enriched cultures, namely 81RN1, OPS, 85RN5, 89SR3-2 and 112YL1 were found to give high cumulative hydrogen formation of 1085, 1009, 994, 983 and 778 mL H₂/L-OP sap, respectively, with the hydrogen content of 29.8, 29.4, 28.7, 27.1 and 27.5%, respectively. PCR–DGGE profiling showed that all these five enriched cultures consisted of species closely related to the genus Clostridium sp. based on the 16S rRNA gene. For pure cultures, the top five hydrogen producers were the isolates encoded as PS-3, PS-4, PS-5, PS-7 and PS-8 exhibiting the hydrogen production of 1973, 1774, 1335, 1170 and 1070 mL H₂/L-OP sap, respectively, with the hydrogen content of 33.7, 29.6, 32.5, 31.5 and 26.4%, respectively. Identification of these high hydrogen producers using 16S rRNA sequence matching showed that the isolates PS-3 and PS-8 belonged to Clostridium beijerinckii, while the isolate PS-7 belonged to Clostridium acetobutylicum and the isolates PS-4 and PS-5 belonged to Klebsiella sp. and Klebsiella pneumoniae, respectively. Therefore, the pure culture C. beijerinckii PS-3 exhibited 1.8 folds higher hydrogen production (1973 mL H₂/L-OP sap) than the enriched cultures of 81RN1 (1085 mL H₂/L-OP sap).     

Fermentative hydrogen production by new marine Clostridium amygdalinum strain C9 isolated from offshore crude oil pipeline

The present study investigated hydrogen production potential of novel marine Clostridium amygdalinum strain C9 isolated from oil water mixtures. Batch fermentations were carried out to determine the optimal conditions for the maximum hydrogen production on xylan, xylose, arabinose and starch. Maximum hydrogen production was pH and substrate dependant. The strain C9 favored optimum pH 7.5 (40 mmol H₂/g xylan) from xylan, pH 7.5–8.5 from xylose (2.2–2.5 mol H₂/mol xylose), pH 8.5 from arabinose (1.78 mol H₂/mol arabinose) and pH 7.5 from starch (390 ml H₂/g starch). But the strain C9 exhibited mixed type fermentation was exhibited during xylose fermentation. NaCl is required for the growth and hydrogen production. Distribution of volatile fatty acids was initial pH dependant and substrate dependant. Optimum NaCl requirement for maximum hydrogen production is substrate dependant (10 g NaCl/L for xylose and arabinose, and 7.5 g NaCl/L for xylan and starch).     

Cell growth and hydrogen production on the mixture of xylose and glucose using a novel strain of Clostridium sp. HR-1 isolated from cow dung compost

A novel mesophilic hydrogen-producing bacterium was isolated from cow dung compost and designated as Clostridium sp. HR-1 by 16S rRNA gene sequence. The optimum condition for hydrogen production by strain HR-1 was pH of 6.5, temperature of 37 °C and yeast extract as nitrogen sources. The strain HR-1 has the ability to utilize kinds of hexose and pentose as carbon sources for growth and H₂ production. Cell growth and hydrogen productivity were investigated for batch fermentation on media containing different ratios of xylose and glucose. Glucose was the preferred substrate in the glucose and xylose mixtures. The high glucose fraction had higher cell biomass production rate. The rate of glucose consumption was higher than xylose consumption, and remained essentially constant independent of xylose content of the mixture. The rate of xylose utilization was decreased with increasing of the glucose fraction. The average H₂ yield and specific H₂ production rates with xylose and glucose are 1.63 mol-H₂/mol xylose and 11.14-H₂ mmol/h g-cdw, and 2.02 mol-H₂/mol-glucose and 9.37 mmol-H₂/h g-cdw, respectively. Using the same initial substrate concentration, the maximum average H₂ yield and specific H₂ production rates with the mixtures of 9 g/l xylose and 3 g/l glucose was 2.01 mol-H₂/mol-mixed sugar and 12.56 mmol-H₂/h g-cdw, respectively. During the fermentation, the main soluble microbial products were ethanol and acetate which showed trends with the different ratios of xylose and glucose.     

Hydrogen production characteristics from dark fermentation of maltose by an isolated strain F.P 01

A hydrogen producing strain F.P 01 was newly isolated from cow dung sludge in an anaerobic bioreactor. The strain F.P 01 was a mesophilic and facultative anaerobic bacterium, which exhibited gram-negative staining in both the exponential and stationary growth phases, and a regular long rod-shaped bacteria with the size of 0.6–0.9 μm × 1.2–2.5 μm, and also could biodegrade a variety of carbohydrates such as glucose, xylose, maltose, etc. The effects of important process parameters on hydrogen producing of F.P 01 were further investigated from hydrogen fermentation of maltose by strain F.P 01, including substrate concentration, medium pH, etc. And the results showed that hydrogen production potential and hydrogen production rate from maltose of this strain F.P 01 was180 mLH₂/g-maltose and 4.0 mLH₂/h, respectively. The corresponding hydrogen concentration of 58–73% was also be observed. Both butyric acid and acetic acid as main by-product was left in the reactor.     

Co-fermentation of a mixture of glucose and xylose to hydrogen by Thermoanaerobacter thermosaccharolyticum W16: Characteristics and kinetics

Glucose and xylose co-fermentation is crucial to maximize hydrogen yield from waste lignocellulose. In this study, cell growth, sugar consumption, and hydrogen production profiles of Thermoanaerobacter thermosaccharolyticum W16 feeding with a range of glucose and xylose were experimental investigated coupled with kinetic analysis. Results showed although T. thermosaccharolyticum W16 could use both glucose and xylose for hydrogen production, a maximum cell growth rate of 0.27 g/L/h and hydrogen production rate of 14.53 mmol/L/h was found with glucose as sole substrate, the value was 92.8% and 49.8% higher than using xylose as the only carbon source. Further interpolation analysis and experimental demonstration suggested when glucose content in the mixed substrate higher than 58.2%, the inhibitory effect on xylose utilization was increased, but when glucose concentration fell below 21.7%, its utilization will be subject to a certain degree of feedback inhibition. Coupling experimental results with kinetic analysis in this study provides a powerful evidence to further develop the potential of T. thermosaccharolyticum W16 as a biocatalyst for hydrogen production from lignocellulosic biomass.     

Microbial hydrogen production from sewage sludge bioaugmented with a constructed microbial consortium

A constructed microbial consortium was formulated from three facultative H₂-producing anaerobic bacteria, Enterobacter cloacae IIT-BT 08, Citrobacter freundii IIT-BT L139 and Bacillus coagulans IIT-BT S1. This consortium was tested as the seed culture for H₂ production. In the initial studies with defined medium (MYG), E. cloacae produced more H₂ than the other two strains and it also was found to be the dominant member when consortium was used. On the other hand, B. coagulans as a pure culture gave better H₂ yield (37.16 ml H₂/g COD_consumed) than the other two strains using sewage sludge as substrate. The pretreatment of sludge included sterilization (15% v/v), dilution and supplementation with 0.5% w/v glucose, which was found to be essential to screen out the H₂ consuming bacteria and ameliorate the H₂ production. Considering (1:1:1) defined consortium as inoculum, COD reduction was higher and yield of H₂ was recorded to be 41.23 ml H₂/g COD_reduced. Microbial profiling of the spent sludge showed that B. coagulans was the dominant member in the constructed consortium contributing towards H₂ production. Increase in H₂ yield indicated that in consortium, the substrate utilization was significantly higher. The H₂ yield from pretreated sludge (35.54 ml H₂/g sludge) was comparatively higher than that reported in literature (8.1–16.9 ml H₂/g sludge). Employing formulated microbial consortium for biohydrogen production is a successful attempt to augment the H₂ yield from sewage sludge.     

Enhanced hydrogen uptake/release in 2LiH–MgB2 composite with titanium additives

The influence of different titanium additives on hydrogen sorption in LiH–MgB₂ system has been investigated. For all the composites LiH–MgB₂–X (X = TiF₄, TiO₂, TiN, and TiC), prepared by ball-milling in molar ratios 2:1:0.1, five hydrogen uptake/release cycles were performed. In-situ synchrotron radiation powder X-ray diffraction (SR-PXD) and attenuated total reflection infrared spectroscopy (ATR-IR) have been used to characterize crystal phases developed during the hydrogen absorption–desorption cycles.     

Thermophilic fermentative hydrogen production from various carbon sources by anaerobic mixed cultures

In the present work, various carbon sources, xylose, glucose, galactose, sucrose, cellobiose, and starch were tested for thermophilic (60 °C) fermentative hydrogen production (FHP) by using the anaerobic mixed culture. An inoculum was obtained from a continuously-stirred tank reactor (CSTR) operated at pH 5.5 and HRT 12 h, and fed with tofu processing waste. The dominant species in the CSTR were found to be Thermoanaerobacterium thermosaccharolyticum and Clostridium thermosaccharolyticum, which are well known thermophilic H₂-producers in anaerobic-state, and have the ability to utilize a wide range of carbohydrates. When initial pH was adjusted to 6.8 ± 0.1 but not controlled during fermentation, vigorous pH drop began within 5 h, and finally reached 4.0–4.5 in all carbon sources. Although over 90% of substrate removal was achieved for all carbon sources except cellobiose (71.7%), the fermentation performances were profoundly different with each other. Glucose, galactose, and sucrose exhibited relatively higher H₂ yields whereas lower H₂ yields were observed for xylose, cellobiose, and starch. On the other hand, when pH was controlled (pH ≥ 5.5), the fermentation performance was enhanced in all carbon sources but to a different extent. A substantial increase in H₂ production was observed for cellobiose, a 1.9-fold increase of H₂ yield along with a substrate removal increase to 93.8%, but a negligible increase for xylose. H₂ production capabilities of all carbon sources tested were as follows: sucrose > galactose > glucose > cellobiose > starch > xylose. The maximum H₂ yield of 3.17 mol H₂/mol hexose_added achieved from sucrose is equivalent to a 26.5% conversion of energy content in sucrose to H₂. Acetic and butyric acids were the main liquid-state metabolites of all carbon sources while lactic acid was detected only in cellobiose, starch and xylose exhibiting relatively lower H₂ yields.     

Improved hydrogen storage properties of LiBH4 doped Li–N–H system

Remarkable improvement of hydrogen sorption properties of Li–N–H system has been obtained by doping with a small amount of LiBH₄. The starting and ending temperatures of hydrogen desorption shift to lower temperatures and the release of NH₃ is obviously restrained by 10 mol% LiBH₄ doping. The kinetics of hydrogen desorption and absorption of Li–N–H system became faster by the addition of LiBH₄. About 4 wt.% H₂ can be released within 30 min and ∼4.8 wt.% H₂ can be reabsorbed within 2 min by LiBH₄ doped sample at 250 °C, while only 1.44 wt.% H₂ is released and 2.1 wt.% is reabsorbed for pure Li–N–H system. The quaternary hydride (LiNH₂)_x(LiBH₄)_(1−x) formed by the reaction between LiBH₄ and LiNH₂ may contribute to the enhancement of the hydrogen sorption performances by yielding a ionic liquid phase and transferring LiNH₂ from solid state to molten state with a weakened N–H bond.