Four products from Escherichia coli pseudogenes increase hydrogen production

Pseudogenes are considered to be nonfunctional genes that lack a physiological role. By screening 3985 Escherichia coli mutants using chemochromic membranes, we found four pseudogenes involved in hydrogen metabolism. Knockouts of pseudogenes ydfW and ypdJ had a defective hydrogen phenotype on glucose and formate, respectively. Also, the knockout of pseudogene yqiG formed hydrogen from formate but not from glucose. For the yqiG mutant, 100% hydrogen recovery was obtained by the complementation of YqiG via a plasmid. The knockout of pseudogene ylcE showed hydrogen deficiency in minimal media which suggested that the role of YlcE is associated with cell growth. Hence, the products of these four pseudogenes play an important physiological role in hydrogen production in E. coli.     

Feasibility of biohydrogen production from industrial wastes using defined microbial co-culture

Background

The development of clean or novel alternative energy has become a global trend that will shape the future of energy. In the present study, 3 microbial strains with different oxygen requirements, including Clostridium acetobutylicum ATCC 824, Enterobacter cloacae ATCC 13047 and Kluyveromyces marxianus 15D, were used to construct a hydrogen production system that was composed of a mixed aerobic-facultative anaerobic-anaerobic consortium. The effects of metal ions, organic acids and carbohydrate substrates on this system were analyzed and compared using electrochemical and kinetic assays. It was then tested using small-scale experiments to evaluate its ability to convert starch in 5 L of organic wastewater into hydrogen. For the one-step biohydrogen production experiment, H1 medium (nutrient broth and potato dextrose broth) was mixed directly with GAM broth to generate H2 medium (H1 medium and GAM broth). Finally, Clostridium acetobutylicum ATCC 824, Enterobacter cloacae ATCC 13047 and Kluyveromyces marxianus 15D of three species microbial co-culture to produce hydrogen under anaerobic conditions. For the two-step biohydrogen production experiment, the H1 medium, after cultured the microbial strains Enterobacter cloacae ATCC 13047 and Kluyveromyces marxianus 15D, was centrifuged to remove the microbial cells and then mixed with GAM broth (H2 medium). Afterward, the bacterial strain Clostridium acetobutylicum ATCC 824 was inoculated into the H2 medium to produce hydrogen by anaerobic fermentation.

Results

The experimental results demonstrated that the optimum conditions for the small-scale fermentative hydrogen production system were at pH 7.0, 35°C, a mixed medium, including H1 medium and H2 medium with 0.50 mol/L ferrous chloride, 0.50 mol/L magnesium sulfate, 0.50 mol/L potassium chloride, 1% w/v citric acid, 5% w/v fructose and 5% w/v glucose. The overall hydrogen production efficiency in the shake flask fermentation group was 33.7 mL/h^-1.L^-1, and those the two-step and the one-step processes of the small-scale fermentative hydrogen production system were 41.2 mL/h^-1.L^-1 and 35.1 mL/h^-1.L^-1, respectively.

Conclusion

Therefore, the results indicate that the hydrogen production efficiency of the two-step process is higher than that of the one-step process.     

Biohydrogen from thermophilic co-fermentation of swine manure with fruit and vegetable waste: maximizing stable production without pH control

Hydrogen production by dark fermentation may suffer of inhibition or instability due to pH deviations from optimality. The co-fermentation of promptly degradable feedstock with alkali-rich materials, such as livestock wastes, may represent a feasible and easy to implement approach to avoid external adjustments of pH.Experiments were designed to investigate the effect of the mixing ratio of fruit-vegetable waste with swine manure with the aim of maximizing biohydrogen production while obtaining process stability through the endogenous alkalinity of manure.Fruit-vegetable/swine manure ratio of 35/65 and HRT of 2 d resulted to give the highest production rate of 3.27 ± 0.51 L_H2 L⁻¹ d⁻¹, with a corresponding hydrogen yield of 126 ± 22 mL_H2 g⁻¹_VS-added and H₂ content in the biogas of 42 ± 5%. At these operating conditions the process exhibited also one of the highest measured stability, with daily productions deviating for less than 14% from the average.     

Optimization of two-phase thermophilic anaerobic digestion of biowaste for hydrogen and methane production through reject water recirculation

The optimization of a two-phase thermophilic anaerobic process treating biowaste for hydrogen and methane production was carried out at pilot scale using two stirred reactors (CSTRs) and without any physical/chemical pre-treatment of inoculum. During the experiment the hydrogen production at low hydraulic retention time (3d) was tested, both with and without reject water recirculation and at two organic loading rate (16 and 21 kgTVS/m³d). The better yields were obtained with recirculation where the pH reached an optimal value (5.5) thanks to the buffering capacity of the recycle stream. The specific gas production of the first reactor was 51 l/kgVS_fed and H₂ content in biogas 37%. The mixture of gas obtained from the two reactors met the standards for the biohythane mix only when lower loading rate were applied to the first reactor, with a composition of 6.7% H₂, 40.1% CO₂ and 52.3% CH₄ the overall SGP being 0.78 m³/kgVS_fed.     

Effects of key operational parameters on biohydrogen production via anaerobic fermentation in a sequencing batch reactor

In this study, a series of tests were conducted in a 6 L anaerobic sequencing batch reactor (ASBR) to investigate the effect of pH, hydraulic retention time (HRT) and organic loading rate on biohydrogen production at 28 °C. Sucrose was used as the main substrate to mimic carbohydrate-rich wastewater and inoculum was prepared from anaerobic digested sludge without pretreatment. The reactor was operated initially with nitrogen sparging to form anaerobic condition. Results showed that methanogens were effectively suppressed. The optimum pH value would vary depending on the HRT. Maximum hydrogen production rate and yield of 3.04 L H₂/L reactor d and 2.16 mol H₂/mol hexose respectively were achieved at pH 4.5, HRT 30 h, and OLR 11.0 kg/m³ d. Two relationships involving the propionic acid/acetic acid ratio and ethanol/acetic acid ratio were derived from the analysis of the metabolites of fermentation. Ethanol/acetic acid ratio of 1.25 was found to be a threshold value for higher hydrogen production.     

Enhancement effect of hematite nanoparticles on fermentative hydrogen production

The effects of hematite nanoparticles concentration (0-1600 mg/L) and initial pH (4.0-10.0) on hydrogen production were investigated in batch assays using sucrose-fed anaerobic mixed bacteria at 35 °C. The optimum hematite nanoparticles concentration with an initial pH 8.48 was 200 mg/L, with the maximum hydrogen yield of 3.21 mol H₂/mol sucrose which was 32.64% higher than the blank test. At 200 mg/L hematite nanoparticles concentration, further initial pH optimization experiments indicated that at pH 6.0 the maximum hydrogen yield reached to 3.57 mol H₂/mol sucrose and hydrogen content was 66.1%. The slow release of hematite nanoparticles had been recorded by transmission electron microscopy (TEM). In addition, TEM analysis indicated that the hematite nanoparticles can affect the shape of bacteria, namely, its length increased from ca. 2.0-3.6 μm to ca. 2.6-5.6 μm, and width became narrower.     

Biohydrogen production from lignocellulosic feedstock

Due to the recent energy crisis and rising concern over climate change, the development of clean alternative energy sources is of significant interest. Biohydrogen produced from cellulosic feedstock, such as second generation feedstock (lignocellulosic biomass) and third generation feedstock (carbohydrate-rich microalgae), is a promising candidate as a clean, CO₂-neutral, non-polluting and high efficiency energy carrier to meet the future needs. This article reviews state-of-the-art technology on lignocellulosic biohydrogen production in terms of feedstock pretreatment, saccharification strategy, and fermentation technology. Future developments of integrated biohydrogen processes leading to efficient waste reduction, low CO₂ emission and high overall hydrogen yield is discussed.     

Hydrolysis of lignocellulosic feedstock by novel cellulases originating from Pseudomonas sp. CL3 for fermentative hydrogen production

A newly isolated indigenous bacterium Pseudomonas sp. CL3 was able to produce novel cellulases consisting of endo-β-1,4-d-glucanase (80 and 100 kDa), exo-β-1,4-d-glucanase (55 kDa) and β-1,4-d-glucosidase (65 kDa) characterized by enzyme assay and zymography analysis. In addition, the CL3 strain also produced xylanase with a molecular weight of 20 kDa. The optimal temperature for enzyme activity was 50, 45, 45 and 55 °C for endo-β-1,4-d-glucanase, exo-β-1,4-d-glucanase, β-1,4-d-glucosidase and xylanase, respectively. All the enzymes displayed optimal activity at pH 6.0. The cellulases/xylanase could hydrolyze cellulosic materials very effectively and were thus used to hydrolyze natural agricultural waste (i.e., bagasse) for clean energy (H₂) production by Clostridiumpasteurianum CH4 using separate hydrolysis and fermentation process. The maximum hydrogen production rate and cumulative hydrogen production were 35 ml/L/h and 1420 ml/L, respectively, with a hydrogen yield of around 0.96 mol H₂/mol glucose.     

Econometric models for biohydrogen development

Biohydrogen is considered as an attractive clean energy source due to its high energy content and environmental-friendly conversion. Analyzing various economic scenarios can help decision makers to optimize development strategies for the biohydrogen sector. This study surveys econometric models of biohydrogen development, including input-out models, life-cycle assessment approach, computable general equilibrium models, linear programming models and impact pathway approach. Fundamentals of each model were briefly reviewed to highlight their advantages and disadvantages. The input-output model and the simplified economic input-output life-cycle assessment model proved most suitable for economic analysis of biohydrogen energy development. A sample analysis using input-output model for forecasting biohydrogen development in the United States is given.     

The Interplay of Proton,Electron, and Metabolite Supply for Photosynthetic H2 Production in Chlamydomonas reinhardtii

To obtain a detailed picture of sulfur deprivation-induced H₂ production in microalgae, metabolome analyses were performed during key time points of the anaerobic H₂ production process of Chlamydomonas reinhardtii. Analyses were performed using gas chromatography coupled to mass spectrometry (GC/MS), two-dimensional gas chromatography combined with time-of-flight mass spectrometry (GCxGC-TOFMS), lipid and starch analysis, and enzymatic determination of fermentative products. The studies were designed to provide a detailed metabolite profile of the solar Bio-H₂ production process. This work reports on the differential analysis of metabolic profiles of the high H₂-producing strain Stm6Glc4 and the wild-type cc406 (WT) before and during the H₂ production phase. Using GCxGC-TOFMS analysis the number of detected peaks increased from 128 peaks, previously detected by GC/MS techniques, to ∼1168. More detailed analysis of the anaerobic H₂ production phase revealed remarkable differences between wild-type and mutant cells in a number of metabolic pathways. Under these physiological conditions the WT produced up to 2.6 times more fatty acids, 2.2 times more neutral lipids, and up to 4 times more fermentation products compared with Stm6Glc4. Based on these results, specific metabolic pathways involving the synthesis of fatty acids, neutral lipids, and fermentation products during anaerobiosis in C. reinhardtii have been identified as potential targets for metabolic engineering to further enhance substrate supply for the hydrogenase(s) in the chloroplast.