Control strategies for hydrogen production through co-culture of Ethanoligenens harbinense B49 and immobilized Rhodopseudomonas faecalis RLD-53

This study evaluated hydrogen production by co-culture of Ethanoligenens harbinense B49 and immobilized Rhodopseudomonas faecalis RLD-53 with different control strategies. To enhance cooperation of dark and photo-fermentation bacteria during hydrogen production process, the glucose concentration, phosphate buffer concentration and initial pH were controlled at 6 g/l, 50 mmol/l and 7.5, respectively. The maximum yield and rate of hydrogen production were 3.10 mol H₂/mol glucose and 17.2 mmol H₂/l/h, respectively. Ethanol from E. harbinense B49 in acetate medium can enhance hydrogen production by R. faecalis RLD-53 except the ratio of ethanol to acetate (R_E/A) among 0.8 to 1.0. Control of the proper phosphate buffer concentration (50 mmol/l) not only increased acetic acid production by E. harbinense B49, but also maintained stable pH of co-culture system. Therefore, the results showed that co-culture of E. harbinense B49 and immobilized R. faecalis RLD-53 was a promising way of converting glucose into hydrogen.     

Alkalinity and high total solids affecting H2 production from organic solid waste by anaerobic consortia

The optimization of total solids in the feed (%TS) and alkalinity ratio (γ) for H₂ production from organic solid wastes under thermophilic regime was carried out using response surface methodology based on a central composite design. The total solids levels were 20.9, 23.0, 28.0, 33.0 and 35.1% whereas the levels of alkalinity ratio (defined as g phosphate alkalinity/g dry substrate) were 0.15, 0.20, 0.30, 0.41 and 0.45. High levels of TS and γ affected in a negative way the H₂ productivity and yield; both response variables significantly increased upon decreasing the TS content and alkalinity ratio. The highest H₂ productivity and yield were 463.7 N mL/kg-d and 54.8 N mL/g VS_rem, respectively, predicted at 20.9% TS and alkalinity ratio 0.25 (0.11 g CaCO₃/g dry substrate). The alkalinity requirements for hydrogenogenic processes were lower than those reported for methanogenic processes (0.11 vs. 0.30 g CaCO₃/g COD). Adequate alkalinity ratio was necessary to maintain optimal biological activity for hydrogen production; however, excessive alkalinity negatively affected process performance probably due to an increase of osmotic pressure. Interestingly, reactor pH depended only on the alkalinity ratio, thus the buffer capacity was able to maintain a constant pH independently of TS levels. At γ = 0.15–0.30 the pHs were in the range 5.56–5.95, which corresponded to the highest hydrogen productivities and yields. Finally, the highest metabolite accumulation corresponded with the highest removal efficiencies but not with high H₂ productivities and yields. Therefore, it seems that organic matter removal was channeled toward solvent generation instead of hydrogen production at high TS and γ levels. This is the first study that shows the requirements of alkalinity in solid substrate fermentation conditions for H₂ production processes and their interaction with the content of total solids in the feed.     

Hydrogen generation from glycerol in batch fermentation process

The influence of concentration of glycerol, inoculum and total nitrogen on hydrogen generation, in batch dark fermentation process in the presence of digested sludge (at 37 °C and at initial pH = 6) was studied. Changes in substrate and products concentrations were modeled with modified Gompertz equations (correlation coefficient R² = 0.9015). The 1,3-propandiol, butyric acid, acetic acid, lactic acid and ethanol were found as the main liquid metabolites. Maximal substrate yield for hydrogen was 0.41 mol H₂/mol glycerol and was obtained for medium containing 10 g/l of glycerol with the lowest amount of inoculum – 1.16 g volatile suspended solid (VSS)/l. Increase of glycerol concentration from 5 to 30 g/l resulted in much better hydrogen generation, namely from 0.345 to 0.715 l H₂/l. Further increase of glycerol concentration did not cause any changes. The H₂:CO₂ ratio in biogas in system with the highest substrate yield was always 1. The initial concentration of glycerol does not influence the rate of hydrogen generation. The increase of initial concentration of inoculum from 1.2 to 11.6 g VSS/l results in the decrease of specific hydrogen yield. Nitrogen concentration in medium does not influence the hydrogen production.     

Inhibitory effects of acetate and ethanol on biohydrogen production of Ethanoligenens harbinese B49

Inhibitory effects of acetate and ethanol on biohydrogen production from glucose by Ethanoligenens harbinese B49 were investigated in this study. In batch test, sodium acetate (0, 10, 20, 50, 100 and 150 mmol/l) and ethanol (0, 20, 40, 80, 100 and 200 mmol/l) were added respectively. Their inhibitory effects on glucose degradation, cell growth, distribution of liquid products and hydrogen production were discussed. Compared with ethanol, acetate exhibited more significant inhibition on growth and hydrogen producing performance of E. harbinese B49. The inhibitory effects of acetate and ethanol were compared and analyzed on the basis of a noncompetitive product inhibition model. For acetate addition, the maximum specific hydrogen production rate r_max = 722 ml/gVSS/h, inhibition constant K_C = 55 mmol/l and the exponent of inhibition n = 0.6 were estimated, whereas the maximum hydrogen yield r_max = 2.2 mol H₂/mol glucose, K_C = 57 mmol/l and n = 0.8 were calculated from kinetic analysis. For ethanol addition, the maximum specific rate r_max = 729 ml/g VSS/h, K_C = 139 mmol/l and n = 0.8 were estimated, whereas the maximum hydrogen yield r_max = 2.2 mol H₂/mol glucose, K_C = 153 mmol/l and n = 0.9 were calculated. In addition, deducing from dose-response curves, the C_I,50 values of ethanol and acetate were 154 and 62 mmol/l, respectively. Acetate has a strong inhibitory effect on hydrogen production with ethanol-type fermentation. Thus, hydrogen production can be improved by optimizing the fermentation strategy through removing the acetate as soon as it was produced.     

Biohydrogen using leachate from an industrial waste landfill as inoculum

Since hydrogen is a renewable energy source, biohydrogen has been researched in recent years. However, there is little data on hydrogen fermentation by a leachate from a waste landfill as inoculum. We investigated hydrogen production using a leachate from an industrial waste landfill in Kanagawa prefecture. The results showed no methane gas production and the leachate was a suitable inoculum for hydrogen fermentation. The maximum H₂ yield was 2.67 mol of H₂ per mol of carbohydrate added, obtained at 30 °C and initial pH 7. The acetate and butyrate production was significant when the H₂ yield was higher. The oxidation–reduction potential analysis of the culture suggested that hydrogen-producing bacteria in the leachate were facultatively anaerobic. Scanning electron microscope observations revealed hydrogen-producing bacteria comprised bacilli of about 2 μm in length.     

Phototrophic hydrogen production from acetate and butyrate in wastewater

Phototrophic hydrogen production was conducted using individual substrates, acetate and butyrate, which are the main products of dark fermentation. Effects of initial pH (ranging 5.0–10.0) and individual substrate concentrations (acetate ranging from 800 to 4100 mg/l, and butyrate ranging from 1000 to 5100 mg/l) on phototrophic hydrogen production were evaluated. The maximum hydrogen yields were 2.5 mol-H₂/mol-acetate at an initial pH of 8.0 treating 800 mg/l of acetate, 3.7 mol-H₂/mol-butyrate at an initial pH of 9.0 treating 1000 mg/l of butyrate. Analyses of DGGE (denaturing gradient gel electrophoresis) profiles of 16S rDNA fragments and FISH (fluorescent in situ hybridization) images show that both phototrophic hydrogen-producing sludges comprised only one predominant species resembling Rhodobacter capsulatus with over 80% relative abundance.     

Isolation and characterization of a new hydrogen-producing strain Bacillus sp. FS2011

A new fermentative hydrogen-producing strain FS2011 was isolated from an effluent of bio-hydrogen production reactor, and identified as Bacillus amyloliquefaciens on the basis of 16S rDNA gene sequence. The strain could utilize various carbon and nitrogen sources to produce hydrogen in a broad range of initial pH (5.29–7.38). Phosphate buffer concentration and fermentation temperature significantly affected hydrogen production and cell growth. The maximum hydrogen yield of 2.26 mol/mol was observed at glucose concentration of 10 g/l, beef extract concentration of 2 g/l, initial pH 6.98, phosphate buffer of 20 mmol/l, and 35 °C, indicating FS2011 was a high-efficiency hydrogen-producing bacterium.     

Biohydrogen production by Ethanoligenens harbinense B49: Nutrient optimization

A new hydrogen-producing bacterial strain Ethanoligenens harbinense B49 was examined for its capability of H₂ production with glucose as sole carbon source. The H₂ production was significantly affected by the concentration of the yeast powder and phosphate in the synthetic medium. The optimized concentration of yeast powder was 0.3–0.5 g/L and the maximum hydrogen yield was obtained at the concentration of phosphate about 100–150 mmol/L. The dynamics of hydrogen production showed that rapid evolution of hydrogen appeared to start after the middle-phase of exponential growth (about 8 h). The maximum H₂ yield and specific hydrogen production rate were estimated to be 2.26 mol H₂/mol glucose and 27.74 mmol H₂/g cell, respectively, when 10 g/L of glucose was present in the medium. The possible pathway of hydrogen production by Ethanoligenens sp. B49 during glucose fermentation was oxidative decarboxylation of pyruvate and the NADH pathway.     

Effect of organic loading rate on fermentative hydrogen production from continuous stirred tank and membrane bioreactors

The influence of organic loading rates (OLRs) on the performance of fermentative hydrogen-producing bioreactors operating in continuous stirred tank reactor (CSTR) and membrane bioreactor (MBR) modes was examined. Five OLRs were examined, ranging from 4.0 to 30 g COD L^?1 d^?1, with influent glucose concentrations ranging from 1.3 to 10 g COD L^?1. At OLRs up to 13 g COD L^?1 d^?1, all influent glucose was utilized and the H₂ yield was not significantly influenced by OLR, although the yield in the CSTR mode was significantly higher than that in the MBR mode, 1.25 versus 0.97 mol H₂ (mol Gluc. Conv.)^?1, respectively. At an OLR of 30 g COD L^?1 d^?1, both reactor modes were overloaded with respect to glucose utilization and also had significantly higher H₂ yields of 1.77 and 1.49 mol H₂ (mol Gluc. Conv.)^?1 for the CSTR and MBR modes, respectively, versus the underloaded operation. At the intermediate OLR of 22 g COD L^?1 d^?1, the H₂ yield was maximized at 1.78 mol H₂ (mol Gluc. Conv.)^?1 for both the CSTR and MBR operation. Overall H₂ production was 50% higher in the MBR mode, 0.78 versus 0.51 moles d^?1, because the CSTR mode was overloaded with respect to glucose utilization at this OLR. These results suggest that an optimum OLR that maximizes H₂ yield and H₂ production may be near the OLR that causes overload with respect to substrate utilization. Additionally, while the CSTR mode is easier to operate and provides higher H₂ yields at underloaded and overloaded OLRs, the MBR mode may be preferable when operating near the optimum OLR.     

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.     

Effects of dark/light bacteria ratio on bio-hydrogen production by combined fed-batch fermentation of ground wheat starch

Bio-hydrogen production by combined dark and light fermentation of ground wheat starch was investigated using fed-batch operation. Serum bottles containing heat-treated anaerobic sludge and a mixture of Rhodobacter sp. was fed with a medium containing 20 g dm^?3 wheat powder (WP) at a constant flow rate. The system was operated at different initial dark/light biomass ratios (D/L). The optimum D/L ratio was 1/2 yielding the highest cumulative hydrogen (1548 cm³), yield (65.2 cm³ g^?1 starch), and specific hydrogen production rate (5.18 cm³ g^?1 h^?1). Light fermentation alone yielded higher hydrogen production than dark fermentation due to fermentation of volatile fatty acids (VFAs) to H₂ and CO₂. The lowest hydrogen formation was obtained with D/L ratio of 1/1 due to accumulation of VFAs in the medium.     

Enhanced biohydrogen production from cornstalk wastes with acidification pretreatment by mixed anaerobic cultures

Biohydrogen production from the cornstalk wastes with acidification pretreatment was reported in this paper. Batch tests were carried out to analyze influences of several environmental factors on biohydrogen production from cornstalk wastes. Two predominant bacterial morphologies, namely spore-forming rod shape bacteria and micrococcus were screened, purified, and identified after enriched from a hydrogen-producing fermentor with cow dung composts. The maximum cumulative H₂ yield of 149.69 ml H₂ g⁻¹ TVS was obtained at initial pH 7.0 and substrate concentration 15 g l⁻¹, the value is about 46-fold as compared with that of raw cornstalk wastes. The maximum hydrogen production rate was 7.6 ml H2 h⁻¹. The hydrogen concentration in biogas was 45–56% (v/v) and there was no significant methane observed in the biogas throughout this study. In addition, biodegradation characteristics of the substrate by microorganisms were also discussed. During the conversion of cornstalk wastes into hydrogen, the acetate, propionate, butyrate, and the ethanol were main by-products in the metabolism of hydrogen fermentation. The test results showed that the acidification pretreatment of the substrate plays a crucial role in conversion of the cornstalk wastes into biohydrogen gas by the cow dung composts generating hydrogen.     

Renewable hydrogen generation by steam reforming of glycerol over zirconia promoted ceria supported catalyst

The study focuses on hydrogen production from steam reforming of glycerol over nickel based catalyst promoted by zirconia and supported over ceria. Catalyst was prepared by the wet-impregnation method and characterized by BET surface area analysis, X-ray diffraction technique and scanning electron microscopy (SEM) analysis. The performance of the catalyst was evaluated in terms of hydrogen yield, selectivity and glycerol conversion at 700 °C in a tubular fixed bed reactor. The effect of glycerol concentration in feed, space time (W/F_AO), temperature and time on stream (TOS) was analyzed for the catalyst Ni–ZrO₂/CeO₂ which showed the complete conversion of glycerol and high H₂ yield that corresponds to 3.95 mol of H₂ out of 7 mol. Thermodynamic analysis was also carried out using Aspen HYSYS for system having glycerol concentration 10 wt% and 20 wt% and experimental results were compared with thermodynamics. Kinetic study was carried out for the steam reforming of glycerol over Ni–ZrO₂/CeO₂ catalyst using the power law model. The values of activation energy and order of reaction were found to be 43.4 kJ/mol and 0.3 respectively.     

Development of 10 kW-scale hydrogen generator using chemical hydride

We have developed a hydrogen generator that generates high purity hydrogen gas from the aqueous solution of sodium borohydride, NaBH₄. This paper discussed the performance testing of the hydrogen generator using a Pt-LiCoO₂-coated honeycomb monolith. The NaBH₄ solution hydrolyzed to generate H₂ and sodium metaborate when it contacted the monolith. The gravimetric and the volumetric H₂ densities of the system were 2 wt.% and 1.5 kg H₂/100 l, respectively. The volumetric density was similar to that of the compressed H₂ at 25 MPa. The hydrogen generator successfully provided a maximum H₂ generation rate of 120 nl/min. Assuming a standard PEM (polymer electrolyte fuel cell, PEFC) fuel cell operated at 0.7 V, generating 120 nl/min was equivalent to12 kW.     

Thermodynamic evaluation of methanol steam reforming for hydrogen production

Thermodynamic equilibrium of methanol steam reforming (MeOH SR) was studied by Gibbs free minimization for hydrogen production as a function of steam-to-carbon ratio (S/C = 0–10), reforming temperature (25–1000 °C), pressure (0.5–3 atm), and product species. The chemical species considered were methanol, water, hydrogen, carbon dioxide, carbon monoxide, carbon (graphite), methane, ethane, propane, i-butane, n-butane, ethanol, propanol, i-butanol, n-butanol, and dimethyl ether (DME). Coke-formed and coke-free regions were also determined as a function of S/C ratio.Based upon a compound basis set MeOH, CO₂, CO, H₂ and H₂O, complete conversion of MeOH was attained at S/C = 1 when the temperature was higher than 200 °C at atmospheric pressure. The concentration and yield of hydrogen could be achieved at almost 75% on a dry basis and 100%, respectively. From the reforming efficiency, the operating condition was optimized for the temperature range of 100–225 °C, S/C range of 1.5–3, and pressure at 1 atm. The calculation indicated that the reforming condition required from sufficient CO concentration (<10 ppm) for polymer electrolyte fuel cell application is too severe for the existing catalysts (T_r = 50 °C and S/C = 4–5). Only methane and coke thermodynamically coexist with H₂O, H₂, CO, and CO₂, while C₂H₆, C₃H₈, i-C₄H₁₀, n-C₄H₁₀, CH₃OH, C₂H₅OH, C₃H₇OH, i-C₄H₉OH, n-C₄H₉OH, and C₂H₆O were suppressed at essentially zero. The temperatures for coke-free region decreased with increase in S/C ratios. The impact of pressure was negligible upon the complete conversion of MeOH.     

Photofermentative hydrogen production using purple non-sulfur bacteria Rhodobacter sphaeroides O.U.001 in an annular photobioreactor: A case study

For meeting the increasing demand of energy, biohydrogen production is to be considered in higher yield. Biohydrogen can be produced both by dark and photofermentative process. In this study, the photofermentative pathway is followed by using dl malic acid (IUPAC name: 2-hydroxybutanedioic acid, molecular weight: 134.08744 g mol^?1, molecular formula: C₄H₆O₅) as carbon source. Pure strain of purple non-sulfur (PNS) bacteria: Rhodobacter sphaeroides strain O.U.001 was studied to produce biohydrogen using the photobioreactor. The photobioreactor was constructed aiming the uniform light distribution. The objective of this study was to investigate the performance of 1 L annular photobioreactor operating in indoor conditions. The highest rate of hydrogen production was obtained at 92 h. In the designed photobioreactor, using Rhodobacter sphaeroides strain O.U.001 (initial dl malic acid concentration of 2.01 g L^?1) at an initial pH of 6.8 ± 0.2, temperature 32 ± 2 °C, inoculum volume 10% (v/v), inoculum age of 48 h, 250 rpm (rotation per minute) stirring and light intensity of 15 ± 1.1 W m^?2, the average H₂ production rate was about 6.5 ± 0.1 mL H₂ h^?1 L^?1 media and yield 4.5 ± 0.05 mol of H₂ mol^?1 of dl malic acid. Luedeking–Piret model was applied for the data fitting to determine the relationship between the cell growth and photofermentative hydrogen production. The photofermentative hydrogen production by this PNS bacterium was found to be microbial mixed growth associated function.     

Thermophilic biohydrogen production from energy plants by Caldicellulosiruptor saccharolyticus and comparison with related studies

Air-dried samples of sweet sorghum, sugarcane bagasse, wheat straw, maize leaves and silphium were utilized without chemical pretreatment as sole energy and carbon sources for H₂ production by the extreme thermophilic bacterium Caldicellulosiruptor saccharolyticus. The specific H₂ production rates and yields were determined in the batch fermentation process. The best substrate was wheat straw, with H₂ production capacity of 44.7 L H₂ (kg dry biomass)^?1 and H₂ yield of 3.8 mol H₂ (mol glucose)^?1. Enzymatically pretreated maize leaves exhibited H₂ production of 38 L H₂ (kg dry biomass)^?1. Slightly less H₂ was obtained from homogenized whole plants of sweet sorghum. Sweet sorghum juice was an excellent H₂ source. Silphium trifoliatum was also fermented though with a moderate production. The results showed that drying is a good storage method and raw plant biomass can be utilized efficiently for thermophilic H₂ production. The data were critically compared with recently published observations.     

Optimization of conditions for hydrogen production from brewery wastewater by anaerobic sludge using desirability function approach

Brewery wastewater was converted H₂ by anaerobic sludge in batch experiments. A three-factor three-level experimental design of Box-Behnken method was adopted to find the optimum H₂ production conditions. The effects of three major influence factors, temperature, pH and brewery wastewater concentration (BWC), on H₂ yield and H₂ maximum production rate (R_max) were evaluated by applying response surface methodology (RSM) integrating a desirability function approach. Desirable H₂ yield and R_max simultaneously were achieved under temperature 35.9 °C, pH 5.95 and BWC 6.05 g/l by a desirability function approach which produced the maximum overall desirability 0.894. Correspondingly, the H₂ yield and R_max were 149.6 ml H₂/g COD and 53.6 ml/h, respectively. The verification test confirms that the optimum H₂ yield and R_max measured were in good agreement with the predicted values, suggesting that the desirability function approach with RSM was a useful technique to get the maximum H₂ yield and R_max simultaneously.     

Thermodynamic equilibrium calculations of hydrogen production from the combined processes of dimethyl ether steam reforming and partial oxidation

Thermodynamic analyses of producing a hydrogen-rich fuel-cell feed from the combined processes of dimethyl ether (DME) partial oxidation and steam reforming were investigated as a function of oxygen-to-carbon ratio (0.00–2.80), steam-to-carbon ratio (0.00–4.00), temperature (100 °C–600 °C), pressure (1–5 atm) and product species.Thermodynamically, dimethyl ether processed with air and steam generates hydrogen-rich fuel-cell feeds; however, the hydrogen concentration is less than that for pure DME steam reforming. Results of the thermodynamic processing of dimethyl ether indicate the complete conversion of dimethyl ether to hydrogen, carbon monoxide and carbon dioxide for temperatures greater than 200 °C, oxygen-to-carbon ratios greater than 0.00 and steam-to-carbon ratios greater than 1.25 at atmospheric pressure (P = 1 atm). Increasing the operating pressure has negligible effects on the hydrogen content. Thermodynamically, dimethyl ether can produce concentrations of hydrogen and carbon monoxide of 52% and 2.2%, respectively, at a temperature of 300 °C, and oxygen-to-carbon ratio of 0.40, a pressure of 1 atm and a steam-to-carbon ratio of 1.50. The order of thermodynamically stable products (excluding H₂, CO, CO₂, DME, NH₃ and H₂O) in decreasing mole fraction is methane, ethane, isopropyl alcohol, acetone, n-propanol, ethylene, ethanol and methyl-ethyl ether; trace amounts of formaldehyde, formic acid and methanol are observed.Ammonia and hydrogen cyanide are also thermodynamically favored products. Ammonia is favored at low temperatures in the range of oxygen-to-carbon ratios of 0.40–2.50 regardless of the steam-to-carbon ratio employed. The maximum ammonia content (i.e., 40%) occurs at an oxygen-to-carbon ratio of 0.40, a steam-to-carbon ratio of 1.00 and a temperature of 100 °C. Hydrogen cyanide is favored at high temperatures and low oxygen-to-carbon ratios with a maximum of 3.18% occurring at an oxygen-to-carbon ratio of 0.40 and a steam-to-carbon ratio of 0.00 in the temperature range of 400 °C–500 °C. Increasing the system pressure shifts the equilibrium toward ammonia and hydrogen cyanide.     

Technological advances in hydrogen production by Enterobacter bacteria upon substrate,luminosity and anaerobic conditions

The enhancement of hydrogen production by Enterobacter aerogenes and Enterobacter cloacae from fermentation of carbon sources such as glucose and lactose (from cheese whey permeate) was investigated. Also, the influence of the luminosity (2200 lux) and anaerobic condition (nitrogen and argon gases) were evaluated. The assays were carried out in 50 mL reactors during 108 h. To E. aerogenes/nitrogen/luminosity condition and using glucose as substrate, H₂ production (73.8 mmol/L.d) was higher than using lactose (15.5 mmol/L.d). In the dark fermentation, hydrogen yields were 1.60 mol H₂/mol glucose and 1.36 molH₂/mol lactose. When using E. cloacae, the light fermentation using nitrogen gas resulted in 77 mmol H₂/L.d and 1.62 mol H₂/mol glucose. In addition, for E. cloacae, hydrogen yields using argon gas and luminosity provided 2.39 mol/mol glucose and 2.53 mol/mol lactose. In general, butyric and acetic acid fermentation were observed and favored the target-product (H₂).