Submitted Successfully!
Thank you for your contribution! You can also upload a video entry related to this topic through the link below:
Check Note
Ver. Summary Created by Modification Content Size Created at Operation
1 -- 3035 2022-11-02 21:25:23 |
2 format Meta information modification 3035 2022-11-03 02:28:05 |
Biohydrogen for Sustainable Energy Solutions
Upload a video

Energy plays a crucial role in the sustainable development of modern nations. Hydrogen is considered the most promising alternative fuel as it can be generated from clean and green sources. Moreover, it is an efficient energy carrier because hydrogen burning only generates water as a byproduct. It is generated from natural gas. However, it can be produced using other methods, i.e., physicochemical, thermal, and biological. The biological method is considered more environmentally friendly and pollution free.

energy photofermentation dark fermentation microorganisms biohydrogen
Subjects: Chemistry, Applied
View Times: 89
Revisions: 2 times (View History)
Update Time: 03 Nov 2022
Table of Contents

    1. Feedstock for Biohydrogen Production

    Biological hydrogen production has been carried out using several waste materials and lignocellulosic materials, depending upon their availability and suitability in particular geographic situations. Numerous raw materials such as sugarcane and sugar beet molasses [1][2][3][4][5][6], cheese whey powder [7], coffee drink manufacturing wastewater [8], corn stalk [9], crude glycerol [10], rice slurry [11], starch wastewater [6], paper and pulp industry effluent [12], baggase [13], dairy wastewater [14], vegetable waste [15], palm oil mill waste [16], distillery wastewater, and waste barley [17][18] have been reported.

    2. Diversity of Biohydrogen-Producing Bacteria

    There are numerous types of fermentative hydrogen-producing bacteria. Clostridium sp. is one of the most common anaerobic bacteria. Different species of Clostridium, such as Clostridium butyricum, Clostridium beijerienckii, Clostridium amygdalinum, Clostridium cellolosi, and Clostridium acetobutylicum, have been reported for fermentative hydrogen production [19][20][21][22][23]. Anaerobic bacteria utilize glucose to produce hydrogen, while butyric acid or acetic acid is produced as the product—Chong et al. isolated Clostridium butyricum from POME [24]. The optimum hydrogen production was obtained at pH 5.5.
    Some facultative anaerobic bacteria (e.g., Enterobacter aerogenes) have also been recognized as H2 producers since the hydrogenase enzyme was found in these bacteria [25]. The critical parameters, such as substrate concentration, temperature, pH, inoculum size, and yeast extract, were optimized to obtain a maximum H2 yield of 0.21 L H2 /g glucose. The culture and maintenance of facultative anaerobes are more feasible than obligate anaerobes.
    These microorganisms are further classified into mesophylls and thermophiles based on their growth temperatures. Although thermophiles are cultivated at elevated temperatures, with highly intensive energy requirements, their H2 production can be closer to the theoretical yield than mesophylls by overwhelming the thermodynamic barrier [26][27]. Some photofermentative bacteria require light energy to produce H2 in anoxygenic conditions. Without O2, these photoautotrophs, including cyanobacteria and green algae, produce H2 through biophotolysis using their specific metabolic routes advantageously under defined conditions [26]. Mixed cultures are also considered the best choice for maximum H2 yield. A study by Nicolau shows hydrogen production using heat-treated mesophilic anaerobic sludge inoculum instead of pure culture [28]. The hydrogen yield at pH 5.5 was 0.37 mol H2/mol of carbohydrate, equal to 18.14 L H2/kg of dry solid.

    3. Enzymes

    Life depends on several chemical reactions, most of which are slow. Therefore, enzymes are naturally occurring catalysts to speed up biochemical reactions. The two enzymes involved in hydrogen production are hydrogenase and nitrogenase. The enzyme hydrogenase catalyzes the consumption and generation of H2. After the discovery of this enzyme in 1930 by Stephenson and Stickland, numerous experiments have been conducted to learn more about it. Despite this, its crystal structure was elucidated approximately 20 years ago [29]. It is present in dark fermentative hydrogen-producing bacteria, green algae, and cyanobacteria. The hydrogenase enzyme is classified into three types based on the structure of active sites: NiFe-, Fe-Fe-, and Fe-hydrogenase. The NiFe-hydrogenase is only present in bacteria and archaea, while algae and bacteria have FeFe-hydrogenase. Hence, Fe-hydrogenase is a homodimer and is only present in methanogenic archaea [29][30][31].
    Nitrogenase is another enzyme responsible for the production of H2. It is found in purple non-Sulfur bacteria, archaea, and cyanobacteria [31]. Most atmospheric nitrogen is fixed by cyanobacteria and generates H2 as a byproduct. There are three forms of nitrogenase enzyme: Molybdenum, iron, and vanadium. They are located at the active sites of nitrogen reduction and bind with rare metal centers. Mo-nitrogenase consists of two proteins: dinitrogenase (MoFe protein) and dinitrogenase reductase (Fe protein). The nitrogenase helps to generate ammonium from nitrogen, but in nitrogen-deficient conditions, it starts producing hydrogen in an anaerobic environment [30]. The structure of iron and vanadium nitrogenases are similar to the structure of the Mo form, but they have FeFe and VFe cofactors, respectively. The FeFe and VFe nitrogenases enhance hydrogen production compared to Mo nitrogenase. Only one type of photofermentative bacteria, R. palustris, has been reported to have all three types of these nitrogenases [31].

    4. Factors Affecting the Production of Hydrogen

    4.1. Pretreatment Methods

    The use of food waste and food processing wastewater as feedstock provides several organic compounds and nutrients with enhanced hydrogen production, but some inhibitory compounds affect the production and yield of hydrogen [32]. In addition, different pretreatment methods have been reported to increase the utilization of raw materials for successive hydrogen generation.
    Among the pretreatment methods, hydrolysis and preheating are the most preferred methods. Hydrolysis can be acid/alkaline or ultrasound-assisted. The six-hour alkaline hydrolysis increases H2 generation 206 times at a pH level of 12 [33]. Meanwhile, acid 12 h hydrolysis enhances the production of H2 three-fold at a pH of 2. Hence, the main disadvantages are the utilization of chemicals in large quantities and the requirement of some other processes to neutralize the pH [34]. The ultrasonication of food waste, assisted with hydrolysis, increases H2 yield by 75–88% [35][36][37][38], but investment in equipment and energy cost are the major hurdles to commercializing this method.
    The preheating of food waste is another pretreatment method [39]. The results depicted that, prior to starting fermentation, heating waste for at least 20 min at 90 °C could increase the H2 yield.

    4.2. Effect of Substrate Concentration

    The substrate concentration plays a vital role in H2 production by dark fermentation. When substrate concentration increases, it creates unfavorable conditions and consequently changes the pH, H2 partial pressure, and the concentration of volatile fatty acids. Therefore, substrate inhibition may be minimized by arranging the optimum initial concentration of the substrate [40]. Many researchers have reported inhibition by substrate concentration, but the main focus was on the sources of carbohydrates. The use of wastewater and organic waste as a substrate has rarely been reported in the literature [41]. The fed-batch reactors can be used to avoid substrate inhibition. Some bacteria, such as Enterobacter aerogens, can reduce substrate inhibition by stimulating the microbial activity of H2 production [40]. The effect of substrate concentration on hydrogen production by Lactobacillus casei and Clostridium butyricum was also evaluated [42]. Glucose and galactose were used as carbon sources during the batch process. The results were based on the inoculum utilization of a single species or a mixture. It was observed that L. casei could not utilize galactose properly when used alone, while C. butyricum gave a fast response to galactose usage as a carbon source. On the other hand, the response for glucose utilization was faster by L. casei than C. butyricum under low concentrations of glucose, and, in turn, low hydrogen production was observed because Lactobacillus outcompeted the most significant H2-generating bacteria.

    4.3. Effect of Initial pH

    Initial pH is another essential factor to be considered in the dark fermentation process, and it is noted that each microbe can function effectively in different conditions. For example, the effective pH for hydrogen production is 5–8. When the initial pH becomes lower than 5, hydrogen production reduces by half [43]. A pH range of 5–9 also has been used during the batch fermentation process, but a PH range of 5–6 has been reported as the initial optimum pH [44]. During fermentation, volatile fatty acids reduce the pH of the medium. Therefore, the initial pH was set from 6-7 to compensate for the end of the process [34].
    The effect of pH on hydrogen production by green algae was evaluated [45]. The results showed that the pH of the medium affects the activity of the hydrogenase enzyme. They controlled the pH by adding NaOH and HCl over the range of 6.5–9.0, which does not affect algae growth. It was observed that an increased yield of 2.4% was obtained at a pH of 6.5. A considerable pH value is also required for Purple non-Sulfur bacteria (PNSB) to produce hydrogen via photofermentation. According to studies on hydrogen production during the photo-biological fermentation process, a pH of 7 is best for transporting electrons to the nitrogenase enzymes for H2 generation in the media [46].

    4.4. Effect of Operational Temperature

    The operational temperature significantly affects the production of hydrogen and the activity of enzymes involved in hydrogen generation [47][48]. Thermophilic bacteria observed an enhanced biosynthesis of hydrogen at high temperatures compared to mesophilic bacteria during dark fermentation. The temperature range of 30–55 °C has been reported as optimal for enhanced biohydrogen production [48][49]. Hence, it has been reported that the activity of H2 producers is inhibited at a very extreme temperature of more than 60 °C. Only hyperthermophilic bacteria (Pyrococcus furiosus and Thermotoga maritime) can produce H2 at extreme temperatures. These bacteria can produce H2 at temperatures greater than 80 °C [50].
    PNSBs are also sensitive to different ranges of temperature. For example, a study conducted to show the effect of cultural conditions on H2 production by photofermentation described that the growth rate and the rate of H2 production initially increased with an increase in temperature up to 30 °C. However, after 30 °C, the production rate of H2 gas decreased rapidly because the higher temperature above 30 °C inhibits the activity of the enzyme nitrogenase [46].

    4.5. Effect of Nutrients

    The macronutrients also play a vital role in the growth of bacteria to produce H2. The essential nutrients are sulfur, phosphorus, and nitrogen. The common form of inorganic sulfur in many organic wastes is sulfate (SO42−) [51]. The sulfate-reducing bacteria reduce sulfate into sulfide during the process of fermentation. The sulfur-containing proteins also produce sulfide in the fermentation medium. Studies have been reported about the toxic effects of high sulfide levels in a medium, which inhibit the activity of microorganisms from producing H2. The increased sulfide concentration also decreases the bioavailability of some trace elements [52].
    Another essential nutrient for the growth of anaerobic bacteria is nitrogen. The high concentration of ammonia hydrogen decreases the activity of fermentative bacteria and the rate of H2 production [53][54]. The degradation of proteins and amino acids also produces a high amount of ammonia in the fermentation media. The high concentration of nitrogen also interferes with the intracellular pH and affects the performance of microbes responsible for H2 production. The inhibition of nitrogen can be overcome by diluting the feedstock [55]. Besides nitrogen and sulfur, phosphorus is another nutrient required to enhance hydrogen production [56]. It was observed that a high rate of H2 can be obtained in the presence of 600 mg L−1 K2HPO4 [57]. A 40% increase in the production of H2 was observed at a 30% increase or decline in the respective chemical compound.

    4.6. Effect of Light Intensity

    Light intensity plays a significant role in producing H2 by PNSB. It was shown that the performance of PNSB increases with an increase in light intensity from 2500–5000 lx, but a further increase in light intensity reduces the growth and production of hydrogen [58]. It was also observed that the photosynthetic system of PNSB demanded more ATP and reduced power with increasing illumination intensity [58]. The enzyme nitrogenase also requires high ATP to sensitize the cells and produce H2. Hence, the high intensity can become a limiting factor in photohydrogen production.
    Photoinhibition in PNSB was also investigated [59]. They suggested that hydrogen production decreased when light intensity was increased above 200 Wm−2, while a study conducted by Cai and Wang found that the H2 production decreased at an illumination intensity of 6000 lx. The favorable light source is LED because it has a wavelength range of 770–920 nm, which is considered best for the activity of PNSB. Furthermore, LED light is cost-effective in terms of heat generation, energy consumption, and life expectancy [60].

    4.7. Effect of Metal Ions

    Different metal ions are used for microbes’ growth and to optimize the activity of the enzyme during dark- and photofermentation. These metal ions are required only in a moderate amount. When they are used in high quantities, they inhibit the fermentation process by inhibiting the growth of the bacteria. The effects of using higher concentrations of metals include destroying membrane function and eliminating the transmission of valuable ions and nutrients to the cell and intracellular accumulation of metals [61][62]. A study was conducted to describe the importance of Fe for metabolic changes and its involvement in the expression of non-Fe-S and Fe-S proteins in hydrogenase enzymes. However, when the concentration of Fe increases in the medium, it makes cell clumps and reduces the mass transfer activity. It has been reported that the pure culture requires very little Fe, while mixed culture can tolerate high doses of Fe without an inhibitory effect [44].
    Trace metal ions, such as sodium, magnesium, and calcium, are also needed for the growth of bacteria. High amounts of these trace metals slow down the growth of microbes and become toxic at higher concentrations. The authors of [63] observed a high hydrogen yield in the absence of sodium. The higher concentrations of sodium raise the osmotic pressure, affect the activity of bacteria, and sometimes cause bacterial death. It has been recommended that the sodium concentration should be kept under 20 g L−1 to achieve a maximum level of hydrogen production [64]. Ca2+n is another trace element required for the growth of bacteria and H2 production [65]. Mg2+ is also responsible for cell function and reaction. It is the most demanding ion as a cofactor for 10 types of enzymes involved in the glycolysis process. The Ni2+ has no inhibitory effect on the yield of H2 at a level of 0.1 mg L−1. Hence, no significant measure has been taken to control metal inhibition during fermentation, but a few pretreatment techniques, such as biosorption, electrodialysis, and cofermentation, can effectively overcome metal inhibition problems [66].

    5. Nanotechnology and Biohydrogen

    The vast and newly emerging field of nanotechnology deals with nm-sized particles. The nanoparticles (NPs) have been utilized in several fields, such as biosensors, medicines, immobilization, and the production of biofuels [67][68][69]. The NPs also help produce biohydrogen by influencing the metabolic activities of microbes under aerobic conditions [70]. Nanoparticles prepared by different methods (biological, physical, and chemical) have been reported for H2 production. The NPs of gold, silver, copper, nickel, iron, zinc oxide, palladium, titanium, silica, carbon nanotubes, and activated carbon have been used to enhance H2 production [67][71][72][73]. These nanoparticles provide a larger surface area to adsorb electrons and, hence, enhance the production rate of H2 by stimulating the hydrogen-producing enzymes [74].

    Enhancement of Biohydrogen by Metallic Nanoparticles

    Zhang’s group was the first to use gold nanoparticles to enhance the biosynthesis of H2 [74]. They used artificial wastewater for H2 production via dark fermentation. The preheated and non-heat-treated cultures were used as inoculum. It was observed that the gold nanoparticles successfully increased the metabolic activity of the microbes, enhancing the rate of H2 compared to the control. The cumulative hydrogen yield was maximum when 5 nm gold particles were used [74]. A study evaluating silver NPs for hydrogen production has also been conducted [75]. They used mixed culture and Ag-NPs to produce H2 from glucose. When the concentration of the Ag-NPs was increased up to 20 nM, it affected the activity of the bacteria for enhancing H2 generation. However, there was no increase in hydrogen production rate at more significant concentrations. The higher yield of H2 observed at 20 nM Ag-NPs was 67.6%. The Ag-NPs also increased cell biomass and decreased the lag phase for H2 production.
    Han and colleagues investigated the effect of hematite NPs and initial pH on hydrogen production in mixed bacteria in an anaerobic fed-batch process. The maximum observed H2 yield was 3.21 mol H2/mol−1 sucrose. A transmission electron microscope was used to check the slow discharge of hematite nanoparticles and their effect on the shape of bacteria. Furthermore, a study was conducted utilizing biogenic palladium nanoparticles and palladium ions [71]. The leaf extract of Cortandrum sattvum was used to synthesize palladium nanoparticles. They obtained a maximum H2 yield of 1.48 mol H2/mol−1 glucose using a 5 mg L−1 palladium nanoparticles concentration because of the higher activity of the hydrogenase enzyme. On the other hand, palladium ions showed a negative impact on the yield and lag phase of hydrogen.
    Many bacterial cultures have been investigated for producing H2 via iron NPs. For example, Fe-NPs and iron ions were used to investigate their possible enhancement effect on the production of H2 [76]. Both showed a positive impact on the hydrogen yield compared to the control. However, Fe+2 ions and Fe-NPs illustrated different behavior towards the generation of intermediate metabolites. The propionate production declined by 75% with Fe-NPs compared to a 35% reduction by the Fe+2 ions. The enhancement effect of phytogenic iron nanoparticles and iron ions was also investigated. The green Fe-NPs were prepared using the extract of leaves and bark of Syzygium cumini and FeSO4. The mesophilic bacterial strain of Enterobacter cloacae DH-89 was isolated from the soil and used to produce hydrogen. A 100% increase in hydrogen production (1.9 mol H2/mol−1 hexose) was observed under 100 mg L−1 Fe-NPs compared to the control (0.95 mol H2/mol −1 glucose). Meanwhile, Fe+2 ions helped to raise the yield of hydrogen to 1.45 mol/mol−1 glucose) [77]. Similarly, some other researchers have also reported Fe, Fe2O3, and Fe3O4 NPs prepared by different physical, chemical, and biological methods for the enhanced biosynthesis of hydrogen [20][78][79][80][81][82][83].
    The effect of ZnO nanoparticles on hydrogen production was also reported [84]. The ZnO-NPs were synthesized by the typical precipitation method. The pretreated biomass of water hyacinth was saccharified by the enzyme activity and used for the fermentative production of H2. It was observed that the ZnO-NPs reduced the hydrogen yield compared to the control. On the other hand, metallic NPs (copper, nickel, silicon dioxide, and titanium dioxide) positively affected the rate of generation and yield of H2 [85][86][87][88][89].
    Many studies have reported an enhancement of H2 production via metallic NPs using the dark fermentation process, but few studies have been found in the literature for enhanced photofermentative H2 production by nanoparticles. Zhao et al. investigated the effect of TiO2-NPs on photofermentative H2 production using the effluent of the dark fermentation process as a feedstock [86]. It was observed from the results that the TiO2-NPs enhanced the activity of PNSB for the production of H2 and reduced the activity of the uptake of hydrogenase enzyme. Another study by Pandey et al. reflected a similar enhancement effect of TiO2-NPs for photofermentative H2 production [85]. Meanwhile, Kanwal and colleagues investigated the effect of a phytofabricated nanoscale iron complex for H2 production using photofermentative PNSB [90]. The use of carbon nanotubes (CNTs) has also been reported for improved hydrogen generation by H2-producing bacteria [91].


    1. Li, W.; Cheng, C.; Cao, G.; Ren, N. Enhanced biohydrogen production from sugarcane molasses by adding ginkgo biloba leaves. Bioresour. Technol. 2020, 298, 122523.
    2. Oceguera-Contreras, E.; Aguilar-Juarez, O.; Oseguera-Galindo, D.; Macías-Barragán, J.; Ortiz-Torres, G.; Pita-López, M.L.; Domínguez, J.; Titov, I.; Kamen, A. Establishment of the upstream processing for renewable production of hydrogen using vermicomposting-tea and molasses as substrate. Waste Manag. 2022, 139, 279–289.
    3. Oliveira, C.A.; Fuess, L.T.; Soares, L.A.; Damianovic, M.H.R.Z. Thermophilic biohydrogen production from sugarcane molasses under low ph: Metabolic and microbial aspects. Int. J. Hydrog. Energy 2020, 45, 4182–4192.
    4. Kars, G.; Alparslan, Ü. Valorization of sugar beet molasses for the production of biohydrogen and 5-aminolevulinic acid by rhodobacter sphaeroides o.U.001 in a biorefinery concept. Int. J. Hydrog. Energy 2013, 38, 14488–14494.
    5. Sagir, E.; Ozgur, E.; Gunduz, U.; Eroglu, I.; Yucel, M. Single-stage photofermentative biohydrogen production from sugar beet molasses by different purple non-sulfur bacteria. Bioprocess Biosyst. Eng. 2017, 40, 1589–1601.
    6. Wei, J.; Liu, Z.-T.; Zhang, X. Biohydrogen production from starch wastewater and application in fuel cell. Int. J. Hydrog. Energy 2010, 35, 2949–2952.
    7. Cota-Navarro, C.B.; Carrillo-Reyes, J.; Davila-Vazquez, G.; Alatriste-Mondragón, F.; Razo-Flores, E. Continuous hydrogen and methane production in a two-stage cheese whey fermentation system. Water Sci. Technol. 2011, 64, 367–374.
    8. Jung, K.-W.; Kim, D.-H.; Lee, M.-Y.; Shin, H.-S. Two-stage uasb reactor converting coffee drink manufacturing wastewater to hydrogen and methane. Int. J. Hydrog. Energy 2012, 37, 7473–7481.
    9. Bala-Amutha, K.; Murugesan, A.G. Biohydrogen production using corn stalk employing bacillus licheniformis msu agm 2 strain. Renew. Energy 2013, 50, 621–627.
    10. Chookaew, T.; O-Thong, S.; Prasertsan, P. Fermentative production of hydrogen and soluble metabolites from crude glycerol of biodiesel plant by the newly isolated thermotolerant klebsiella pneumoniae tr17. Int. J. Hydrog. Energy 2012, 37, 13314–13322.
    11. Fang, H.H.P.; Li, C.; Zhang, T. Acidophilic biohydrogen production from rice slurry. Int. J. Hydrog. Energy 2006, 31, 683–692.
    12. Lakshmidevi, R.; Muthukumar, K. Enzymatic saccharification and fermentation of paper and pulp industry effluent for biohydrogen production. Int. J. Hydrog. Energy 2010, 35, 3389–3400.
    13. Anam, K.; Habibi, M.S.; Harwati, T.U.; Susilaningsih, D. Photofermentative hydrogen production using rhodobium marinum from bagasse and soy sauce wastewater. Int. J. Hydrog. Energy 2012, 37, 15436–15442.
    14. Seifert, K.; Waligorska, M.; Laniecki, M. Brewery wastewaters in photobiological hydrogen generation in presence of rhodobacter sphaeroides O.U. 001. Int. J. Hydrog. Energy 2010, 35, 4085–4091.
    15. Adessi, A.; McKinlay, J.B.; Harwood, C.S.; de Philippis, R. A rhodopseudomonas palustris nifa* mutant produces h2 from nh4+-containing vegetable wastes. Int. J. Hydrog. Energy 2012, 37, 15893–15900.
    16. Alam, M.Z.; Jamal, P.; Nadzir, M.M. Bioconversion of palm oil mill effluent for citric acid production: Statistical optimization of fermentation media and time by central composite design. World J. Microbiol. Biotechnol. 2008, 24, 1177–1185.
    17. Akil, K.; Jayanthi, S. The biohydrogen potential of distillery wastewater by dark fermentation in an anaerobic sequencing batch reactor. Int. J. Green Energy 2014, 11, 28–39.
    18. Kars, G.; Ceylan, A. Biohydrogen and 5-aminolevulinic acid production from waste barley by rhodobacter sphaeroides o.U.001 in a biorefinery concept. Int. J. Hydrog. Energy 2013, 38, 5573–5579.
    19. Pattra, S.; Lay, C.-H.; Lin, C.-Y.; O-Thong, S.; Reungsang, A. Performance and population analysis of hydrogen production from sugarcane juice by non-sterile continuous stirred tank reactor augmented with clostridium butyricum. Int. J. Hydrog. Energy 2011, 36, 8697–8703.
    20. Zhao, W.; Zhao, J.; Chen, G.D.; Feng, R.; Yang, J.; Zhao, Y.F.; Wei, Q.; Du, B.; Zhang, Y.F. Anaerobic biohydrogen production by the mixed culture with mesoporous Fe3O4 nanoparticles activation. Presented at Advanced Materials Research. Trans. Tech. Publ. 2011, 306, 1528–1531.
    21. Jayasinghearachchi, H.S.; Singh, S.; Sarma, P.M.; Aginihotri, A.; Lal, B. Fermentative hydrogen production by new marine clostridium amygdalinum strain c9 isolated from offshore crude oil pipeline. Int. J. Hydrog. Energy 2010, 35, 6665–6673.
    22. Cai, J.; Wang, Y.; Liu, J.; Zhang, X.; Li, F. Pretreatment enhanced structural disruption, enzymatic hydrolysis, fermentative hydrogen production from rice straw. Int. J. Hydrog. Energy 2022, 47, 11778–11786.
    23. Zhang, H.; Bruns, M.A.; Logan, B.E. Biological hydrogen production by clostridium acetobutylicum in an unsaturated flow reactor. Water Res. 2006, 40, 728–734.
    24. Chong, M.-L.; Rahim, R.A.; Shirai, Y.; Hassan, M.A. Biohydrogen production by clostridium butyricum eb6 from palm oil mill effluent. Int. J. Hydrog. Energy 2009, 34, 764–771.
    25. Rao, R.; Basak, N. Optimization and modelling of dark fermentative hydrogen production from cheese whey by enterobacter aerogenes 2822. Int. J. Hydrog. Energy 2021, 46, 1777–1800.
    26. Chandrasekhar, K.; Lee, Y.-J.; Lee, D.-W. Biohydrogen production: Strategies to improve process efficiency through microbial routes. Int. J. Mol. Sci. 2015, 16, 8266–8293.
    27. Singh, V.; Das, D. Chapter 3—Potential of hydrogen production from biomass. In Science and Engineering of Hydrogen-Based Energy Technologies; de Miranda, P.E.V., Ed.; Academic Press: Cambridge, MA, USA, 2019; pp. 123–164.
    28. Massanet-Nicolau, J.; Dinsdale, R.; Guwy, A. Hydrogen production from sewage sludge using mixed microflora inoculum: Effect of ph and enzymatic pretreatment. Bioresour. Technol. 2008, 99, 6325–6331.
    29. Xu, T.; Chen, D.; Hu, X. Hydrogen-activating models of hydrogenases. Coord. Chem. Rev. 2015, 303, 32–41.
    30. Khetkorn, W.; Rastogi, R.P.; Incharoensakdi, A.; Lindblad, P.; Madamwar, D.; Pandey, A.; Larroche, C. Microalgal hydrogen production–a review. Bioresour. Technol. 2017, 243, 1194–1206.
    31. Kim, D.-H.; Kim, M.-S. Hydrogenases for biological hydrogen production. Bioresour. Technol. 2011, 102, 8423–8431.
    32. Ghosh, S.; Dairkee, U.K.; Chowdhury, R.; Bhattacharya, P. Hydrogen from food processing wastes via photofermentation using purple non-sulfur bacteria (pnsb)–a review. Energy Convers. Manag. 2017, 141, 299–314.
    33. Jang, S.; Kim, D.-H.; Yun, Y.-M.; Lee, M.-K.; Moon, C.; Kang, W.-S.; Kwak, S.-S.; Kim, M.-S. Hydrogen fermentation of food waste by alkali-shock pretreatment: Microbial community analysis and limitation of continuous operation. Bioresour. Technol. 2015, 186, 215–222.
    34. Jarunglumlert, T.; Prommuak, C.; Putmai, N.; Pavasant, P. Scaling-up bio-hydrogen production from food waste: Feasibilities and challenges. Int. J. Hydrog. Energy 2018, 43, 634–648.
    35. Gadhe, A.; Sonawane, S.S.; Varma, M.N. Ultrasonic pretreatment for an enhancement of biohydrogen production from complex food waste. Int. J. Hydrog. Energy 2014, 39, 7721–7729.
    36. Elbeshbishy, E.; Nakhla, G. Comparative study of the effect of ultrasonication on the anaerobic biodegradability of food waste in single and two-stage systems. Bioresour. Technol. 2011, 102, 6449–6457.
    37. Elbeshbishy, E.; Hafez, H.; Dhar, B.R.; Nakhla, G. Single and combined effect of various pretreatment methods for biohydrogen production from food waste. Int. J. Hydrog. Energy 2011, 36, 11379–11387.
    38. Elbeshbishy, E.; Hafez, H.; Nakhla, G. Viability of ultrasonication of food waste for hydrogen production. Int. J. Hydrog. Energy 2012, 37, 2960–2964.
    39. Kim, D.-H.; Kim, S.-H.; Shin, H.-S. Hydrogen fermentation of food waste without inoculum addition. Enzym. Microb. Technol. 2009, 45, 181–187.
    40. Kothari, R.; Kumar, V.; Pathak, V.V.; Ahmad, S.; Aoyi, O.; Tyagi, V. A critical review on factors influencing fermentative hydrogen production. Front. Biosci. 2017, 22, 1195–1220.
    41. Kim, E.-J.; Kim, M.-S.; Lee, J.K. Hydrogen evolution under photoheterotrophic and dark fermentative conditions by recombinant rhodobacter sphaeroides containing the genes for fermentative pyruvate metabolism of rhodospirillum rubrum. Int. J. Hydrog. Energy 2008, 33, 5131–5136.
    42. Park, J.-H.; Kim, D.-H.; Kim, S.-H.; Yoon, J.-J.; Park, H.-D. Effect of substrate concentration on the competition between clostridium and lactobacillus during biohydrogen production. Int. J. Hydrog. Energy 2017, 43, 11460–11469.
    43. Dong-Jie, N.; Jing-Yuan, W.; Bao-Ying, W.; You-Cai, Z. Effect of mo-containing additives on biohydrogen fermentation from cassava’s stillage. Int. J. Hydrog. Energy 2011, 36, 5289–5295.
    44. Elbeshbishy, E.; Dhar, B.R.; Nakhla, G.; Lee, H.-S. A critical review on inhibition of dark biohydrogen fermentation. Renew. Sustain. Energy Rev. 2017, 79, 656–668.
    45. Maneeruttanarungroj, C.; Phunpruch, S. Effect of ph on biohydrogen production in green alga tetraspora sp. Cu2551. Energy Procedia 2017, 138, 1085–1092.
    46. Wang, B.-N.; Yang, C.-F.; Lee, C.-M. The factors influencing direct photohydrogen production and anaerobic fermentation hydrogen production combination bioreactors. Int. J. Hydrog. Energy 2011, 36, 14069–14077.
    47. Guo, X.M.; Trably, E.; Latrille, E.; Carrere, H.; Steyer, J.-P. Hydrogen production from agricultural waste by dark fermentation: A review. Int. J. Hydrog. Energy 2010, 35, 10660–10673.
    48. Sinha, P.; Pandey, A. An evaluative report and challenges for fermentative biohydrogen production. Int. J. Hydrog. Energy 2011, 36, 7460–7478.
    49. Saady, N.M.C. Homoacetogenesis during hydrogen production by mixed cultures dark fermentation: Unresolved challenge. Int. J. Hydrog. Energy 2013, 38, 13172–13191.
    50. Pawar, S.S.; van Niel, E.W. Thermophilic biohydrogen production: How far are we? Appl. Microbiol. Biotechnol. 2013, 97, 7999–8009.
    51. Hwang, J.-H.; Choi, J.-A.; Oh, Y.-K.; Abou-Shanab, R.A.; Song, H.; Min, B.; Cho, Y.; Na, J.-G.; Koo, J.; Jeon, B.-H. Hydrogen production from sulfate-and ferrous-enriched wastewater. Int. J. Hydrog. Energy 2011, 36, 13984–13990.
    52. Dhar, B.R.; Elbeshbishy, E.; Nakhla, G. Influence of iron on sulfide inhibition in dark biohydrogen fermentation. Bioresour. Technol. 2012, 126, 123–130.
    53. Ahmadi-Pirlou, M.; Ebrahimi-Nik, M.; Khojastehpour, M.; Ebrahimi, S.H. Mesophilic co-digestion of municipal solid waste and sewage sludge: Effect of mixing ratio, total solids, and alkaline pretreatment. Int. Biodeterior. Biodegrad. 2017, 125, 97–104.
    54. del Pilar Anzola-Rojas, M.; da Fonseca, S.G.; da Silva, C.C.; de Oliveira, V.M.; Zaiat, M. The use of the carbon/nitrogen ratio and specific organic loading rate as tools for improving biohydrogen production in fixed-bed reactors. Biotechnol. Rep. 2015, 5, 46–54.
    55. Salerno, M.B.; Park, W.; Zuo, Y.; Logan, B.E. Inhibition of biohydrogen production by ammonia. Water Res. 2006, 40, 1167–1172.
    56. Zhou, P.; Elbeshbishy, E.; Nakhla, G. Optimization of biological hydrogen production for anaerobic co-digestion of food waste and wastewater biosolids. Bioresour. Technol. 2013, 130, 710–718.
    57. Lin, C.; Lay, C. Carbon/nitrogen-ratio effect on fermentative hydrogen production by mixed microflora. Int. J. Hydrog. Energy 2004, 29, 41–45.
    58. Assawamongkholsiri, T.; Reungsang, A. Photo fermentational hydrogen production of rhodobacter sp. Kku-ps1 isolated from an uasb reactor. Electron. J. Biotechnol. 2015, 18, 221–230.
    59. Kim, M.-S.; Baek, J.-S.; Lee, J.K. Comparison of h2 accumulation by rhodobacter sphaeroides kd131 and its uptake hydrogenase and phb synthase deficient mutant. Int. J. Hydrog. Energy 2006, 31, 121–127.
    60. Akroum-Amrouche, D.; Abdi, N.; Lounici, H.; Mameri, N. Effect of physico-chemical parameters on biohydrogen production and growth characteristics by batch culture of rhodobacter sphaeroides cip 60.6. Appl. Energy 2011, 88, 2130–2135.
    61. Lemire, J.A.; Harrison, J.J.; Turner, R.J. Antimicrobial activity of metals: Mechanisms, molecular targets and applications. Nat. Rev. Microbiol. 2013, 11, 371.
    62. Le, D.T.H.; Nitisoravut, R. Modified hydrotalcites for enhancement of biohydrogen production. Int. J. Hydrog. Energy 2015, 40, 12169–12176.
    63. Junghare, M.; Subudhi, S.; Lal, B. Improvement of hydrogen production under decreased partial pressure by newly isolated alkaline tolerant anaerobe, clostridium butyricum tm-9a: Optimization of process parameters. Int. J. Hydrog. Energy 2012, 37, 3160–3168.
    64. Cao, X.; Zhao, Y. The influence of sodium on biohydrogen production from food waste by anaerobic fermentation. J. Mater. Cycles Waste Manag. 2009, 11, 244–250.
    65. Yu, H.-Q.; Tay, J.-H.; Fang, H.H. The roles of calcium in sludge granulation during uasb reactor start-up. Water Res. 2001, 35, 1052–1060.
    66. Singhania, R.R.; Patel, A.K.; Christophe, G.; Fontanille, P.; Larroche, C. Biological upgrading of volatile fatty acids, key intermediates for the valorization of biowaste through dark anaerobic fermentation. Bioresour. Technol. 2013, 145, 166–174.
    67. Elreedy, A.; Ibrahim, E.; Hassan, N.; El-Dissouky, A.; Fujii, M.; Yoshimura, C.; Tawfik, A. Nickel-graphene nanocomposite as a novel supplement for enhancement of biohydrogen production from industrial wastewater containing mono-ethylene glycol. Energy Convers. Manag. 2017, 140, 133–144.
    68. Patel, S.K.; Choi, S.H.; Kang, Y.C.; Lee, J.-K. Large-scale aerosol-assisted synthesis of biofriendly fe 2 o 3 yolk–shell particles: A promising support for enzyme immobilization. Nanoscale 2016, 8, 6728–6738.
    69. Otari, S.; Pawar, S.; Patel, S.K.; Singh, R.K.; Kim, S.-Y.; Lee, J.H.; Zhang, L.; Lee, J.-K. Canna edulis leaf extract-mediated preparation of stabilized silver nanoparticles: Characterization, antimicrobial activity, and toxicity studies. J. Microbiol. Biotechnol. 2017, 27, 731–738.
    70. Beckers, L.; Hiligsmann, S.; Lambert, S.D.; Heinrichs, B.; Thonart, P. Improving effect of metal and oxide nanoparticles encapsulated in porous silica on fermentative biohydrogen production by clostridium butyricum. Bioresour. Technol. 2013, 133, 109–117.
    71. Mohanraj, S.; Anbalagan, K.; Kodhaiyolii, S.; Pugalenthi, V. Comparative evaluation of fermentative hydrogen production using enterobacter cloacae and mixed culture: Effect of Pd(ii) ion and phytogenic palladium nanoparticles. J. Biotechnol. 2014, 192, 87–95.
    72. Mohanraj, S.; Anbalagan, K.; Rajaguru, P.; Pugalenthi, V. Effects of phytogenic copper nanoparticles on fermentative hydrogen production by enterobacter cloacae and clostridium acetobutylicum. Int. J. Hydrog. Energy 2016, 41, 10639–10645.
    73. Gadhe, A.; Sonawane, S.S.; Varma, M.N. Influence of nickel and hematite nanoparticle powder on the production of biohydrogen from complex distillery wastewater in batch fermentation. Int. J. Hydrog. Energy 2015, 40, 10734–10743.
    74. Zhang, Y.; Shen, J. Enhancement effect of gold nanoparticles on biohydrogen production from artificial wastewater. Int. J. Hydrog. Energy 2007, 32, 17–23.
    75. Zhao, W.; Zhang, Y.; Du, B.; Wei, D.; Wei, Q.; Zhao, Y. Enhancement effect of silver nanoparticles on fermentative biohydrogen production using mixed bacteria. Bioresour. Technol. 2013, 142, 240–245.
    76. Taherdanak, M.; Zilouei, H.; Karimi, K. The effects of fe0 and ni0 nanoparticles versus fe2+ and ni2+ ions on dark hydrogen fermentation. Int. J. Hydrog. Energy 2016, 41, 167–173.
    77. Nath, D.; Manhar, A.K.; Gupta, K.; Saikia, D.; Das, S.K.; Mandal, M. Phytosynthesized iron nanoparticles: Effects on fermentative hydrogen production by enterobacter cloacae dh-89. Bull. Mater. Sci. 2015, 38, 1533–1538.
    78. Mahmood, T. Effect of iron nanoparticles on hyacinth’ s fermentation. Int. J. Sci. 2013, 2, 106–121.
    79. Zhang, L.; Zhang, L.; Li, D. Enhanced dark fermentative hydrogen production by zero-valent iron activated carbon micro-electrolysis. Int. J. Hydrog. Energy 2015, 40, 12201–12208.
    80. Lin, R.; Cheng, J.; Ding, L.; Song, W.; Liu, M.; Zhou, J.; Cen, K. Enhanced dark hydrogen fermentation by addition of ferric oxide nanoparticles using enterobacter aerogenes. Bioresour. Technol. 2016, 207, 213–219.
    81. Nasr, M.; Tawfik, A.; Ookawara, S.; Suzuki, M.; Kumari, S.; Bux, F. Continuous biohydrogen production from starch wastewater via sequential dark-photo fermentation with emphasize on maghemite nanoparticles. J. Ind. Eng. Chem. 2015, 21, 500–506.
    82. Reddy, K.; Nasr, M.; Kumari, S.; Kumar, S.; Gupta, S.K.; Enitan, A.M.; Bux, F. Biohydrogen production from sugarcane bagasse hydrolysate: Effects of ph, s/x, Fe2+, and magnetite nanoparticles. Environ. Sci. Pollut. Res. 2017, 24, 8790–8804.
    83. Dolly, S.; Pandey, A.; Pandey, B.K.; Gopal, R. Process parameter optimization and enhancement of photo-biohydrogen production by mixed culture of rhodobacter sphaeroides nmbl-02 and escherichia coli nmbl-04 using fe-nanoparticle. Int. J. Hydrog. Energy 2015, 40, 16010–16020.
    84. Zada, B.; Mahmood, T.; Malik, S.A. Effect of zinc oxide nanoparticles on hyacinth’s fermentation. Int. J. Enhanc. Res. Sci. Technol. Eng. 2014, 3, 78–92.
    85. Pandey, A.; Gupta, K.; Pandey, A. Effect of nanosized tio2 on photofermentation by rhodobacter sphaeroides nmbl-02. Biomass Bioenergy 2015, 72, 273–279.
    86. Zhao, Y.; Chen, Y. Nano-tio2 enhanced photofermentative hydrogen produced from the dark fermentation liquid of waste activated sludge. Environ. Sci. Technol. 2011, 45, 8589–8595.
    87. Giannelli, L.; Torzillo, G. Hydrogen production with the microalga chlamydomonas reinhardtii grown in a compact tubular photobioreactor immersed in a scattering light nanoparticle suspension. Int. J. Hydrog. Energy 2012, 37, 16951–16961.
    88. Mullai, P.; Yogeswari, M.K.; Sridevi, K. Optimisation and enhancement of biohydrogen production using nickel nanoparticles—A novel approach. Bioresour. Technol. 2013, 141, 212–219.
    89. Patel, S.K.S.; Lee, J.-K.; Kalia, V.C. Nanoparticles in biological hydrogen production: An overview. Indian J. Microbiol. 2018, 58, 8–18.
    90. Kanwal, F.; Tahir, A.; Shah, S.A.Q.; Tsuzuki, T.; Nisbet, D.; Chen, J.; Rehman, Y. Effect of phyto-fabricated nanoscale organic-iron complex on photo-fermentative hydrogen production by rhodopseudomonas palustris mp2 and rhodopseudomonas palustris mp4. Biomass Bioenergy 2020, 140, 105667.
    91. Liu, Z.; Lv, F.; Zheng, H.; Zhang, C.; Wei, F.; Xing, X.-H. Enhanced hydrogen production in a uasb reactor by retaining microbial consortium onto carbon nanotubes (cnts). Int. J. Hydrog. Energy 2012, 37, 10619–10626.
    Subjects: Chemistry, Applied
    Contributors MDPI registered users' name will be linked to their SciProfiles pages. To register with us, please refer to : ,
    View Times: 89
    Revisions: 2 times (View History)
    Update Time: 03 Nov 2022
    Table of Contents


      Are you sure to Delete?

      Video Upload Options

      Do you have a full video?
      If you have any further questions, please contact Encyclopedia Editorial Office.
      Kanwal, F.; Torriero, A.A.J. Biohydrogen for Sustainable Energy Solutions. Encyclopedia. Available online: (accessed on 27 November 2022).
      Kanwal F, Torriero AAJ. Biohydrogen for Sustainable Energy Solutions. Encyclopedia. Available at: Accessed November 27, 2022.
      Kanwal, Fariha, Angel A. J. Torriero. "Biohydrogen for Sustainable Energy Solutions," Encyclopedia, (accessed November 27, 2022).
      Kanwal, F., & Torriero, A.A.J. (2022, November 02). Biohydrogen for Sustainable Energy Solutions. In Encyclopedia.
      Kanwal, Fariha and Angel A. J. Torriero. ''Biohydrogen for Sustainable Energy Solutions.'' Encyclopedia. Web. 02 November, 2022.