Applied Microbiology and Biotechnology

, Volume 97, Issue 15, pp 7005–7013 | Cite as

The membraneless bioelectrochemical reactor stimulates hydrogen fermentation by inhibiting methanogenic archaea

  • Kengo Sasaki
  • Masahiko MoritaEmail author
  • Daisuke Sasaki
  • Naoya Ohmura
  • Yasuo Igarashi
Environmental biotechnology


The membraneless bioelectrochemical reactor (Ml-BER) is useful for dark hydrogen fermentation. The effect of the electrochemical reaction on microorganisms in the Ml-BER was investigated using glucose as the substrate and compared with organisms in a membraneless non-bioelectrochemical reactor (Ml-NBER) and bioelectrochemical reactor (BER) with a proton exchange membrane. The potentials on the working electrode of the Ml-BER and BER with membrane were regulated to −0.9 V (versus Ag/AgCl) to avoid water electrolysis with a carbon electrode. The Ml-BER showed suppressed methane production (19.8 ± 9.1 mg-C·L−1·day−1) and increased hydrogen production (12.6 ± 3.1 mg-H·L−1·day−1) at pHout 6.2 ± 0.1, and the major intermediate was butyrate (24.9 ± 2.4 mM), suggesting efficient hydrogen fermentation. In contrast, the Ml-NBER showed high methane production (239.3 ± 17.9 mg-C·L−1·day−1) and low hydrogen production (0.2 ± 0.0 mg-H·L−1·day−1) at pHout 6.3 ± 0.1. In the cathodic chamber of the BER with membrane, methane production was high (276.3 ± 20.4 mg-C·L−1·day−1) (pHout, 7.2 ± 0.1). In the anodic chamber of the BER with membrane (anode-BER), gas production was low because of high lactate production (43.6 ± 1.7 mM) at pHout 5.0 ± 0.1. Methanogenic archaea were not detected in the Ml-BER and anode-BER. However, Methanosarcina sp. and Methanobacterium sp. were found in Ml-NBER. Prokaryotic copy numbers in the Ml-BER and Ml-NBER were similar, as were the bacterial community structures. Thus, the electrochemical reaction in the Ml-BER affected hydrogenotrophic and acetoclastic methanogens, but not the bacterial community.


Hydrogen Fermentation Bioelectrochemical system Microbial community Methanogen 



This research was supported in part by the New Energy and Industrial Technology Development Organization (NEDO), Japan, and a Grant-in-Aid for Young Scientists (B) (24780067). We thank Yumi Kotake for her help.


  1. Abreu AA, Karakashev D, Angelidaki I, Sousa DZ, Alves MM (2012) Biohydrogen production from arabinose and glucose using extreme thermophilic anaerobic mixed cultures. Biotechnol Biofuels 5:6PubMedCrossRefGoogle Scholar
  2. Ben Hania W, Godbane R, Postec A, Hamdi M, Ollivier B, Fardeau ML (2012) Isolation and characterization of Defluviitoga tunisiensis gen. nov, sp. nov., a novel thermophilic bacterium pertaining to the order Thermotogales, isolated from a mesothermic anaerobic reactor treating cheese whey in Tunisia. Int J Syst Evol Microbiol 62(Pt 6):1377–1382. doi: 10.1099/ijs.0.033720-0 PubMedCrossRefGoogle Scholar
  3. Bryant MP, Boone DR (1987) Isolation and characterization of Methanobacterium formicicum MF. Int J Syst Bacteriol 37:171CrossRefGoogle Scholar
  4. Cardinali-Rezende J, Debarry RB, Colturato LFDB, Carneiro EV, Chartone-Souza E, Nascimento AMA (2009) Molecular identification and dynamics of microbial communities in reactor treating organic household waste. Appl Microbiol Biotechnol 84:777–789PubMedCrossRefGoogle Scholar
  5. Chong ML, Rahman NAA, Yee PL, Aziz SA, Rahim RA, Shirai Y, Hassan MA (2009) Effect of pH, glucose and iron sulfate concentration on the yield of biohydrogen by Clostridium butyricum EB6. Int J Hydrog Energy 34:8859–8865CrossRefGoogle Scholar
  6. Cirne DG, Bond P, Pratt S, Lant P, Batstone DJ (2012) Microbial community analysis during continuous fermentation of thermally hydrolysed waste activated sludge. Water Sci Technol 65:7–14PubMedGoogle Scholar
  7. Clauwaert P, Verstraete W (2009) Methanogenesis in membraneless microbial electrolysis cells. Appl Microbiol Biotechnol 82:829–836PubMedCrossRefGoogle Scholar
  8. Freguia S, Rabaey K, Yuan Z, Kerrer J (2008) Syntrophic processed drive the conversion of glucose in microbial fuel cell anodes. Environ Sci Technol 42:7937–7943PubMedCrossRefGoogle Scholar
  9. Harold FM, Levin E (1974) Lactic acid translocation: terminal step in glycolysis by Streptococcus faecalis. J Bacteriol 117:1141–1148PubMedGoogle Scholar
  10. Hawkes FR, Hussy I, Kyazze G, Dinsdale R, Hawkes DL (2007) Continuous dark fermentative hydrogen production by mesophilic microflora: principles and progress. Int J Hydrog Energy 32:172–184CrossRefGoogle Scholar
  11. Hoover SR, Porges N (1952) Assimilation of dairy wastes by activated sludge. II. The equation of synthesis and oxygen utilization. Sewage Ind Wastes 24:306–312Google Scholar
  12. Jung S, Regan JM (2011) Influence of external resistance on electrogenesis, methanogenesis, and anode prokaryotic communities in microbial fuel cells. Appl Environ Microbiol 77:564–571PubMedCrossRefGoogle Scholar
  13. Jungermann K, Thauer RK, Leimenstoll G, Decker K (1973) Function of reduced pyridine nucleotide-ferredoxin oxidoreductases in saccharolytic Clostridia. Biochim Biophys Acta 305:268–280PubMedCrossRefGoogle Scholar
  14. Lane DJ (1991) 16S/23S rRNA sequencing. In: Stackebrandt E, Goodfellow M (eds) Nucleic acid techniques in bacterial systematics. Wiley, New York, pp 115–175Google Scholar
  15. Lee HS, Vermaas WFJ, Rittmann BE (2010) Biological hydrogen production: prospects and challenges. Trends Biotechnol 28:262–271PubMedCrossRefGoogle Scholar
  16. Li C, Fang HHP (2007) Fermentative hydrogen production from wastewater and solid wastes by mixed cultures. Crit Rev Environ Sci Technol 37:1–39CrossRefGoogle Scholar
  17. Logan BE (2010) Scaling up microbial fuel cells and other bioelectrochemical systems. Appl Microbiol Biotechnol 85:1665–1671PubMedCrossRefGoogle Scholar
  18. Lueders T, Friedrich MW (2002) Effects of amendment with ferrihydrite and gypsum on the structure and activity of methanogenic populations in rice field soil. Appl Environ Microbiol 68:2484–2494PubMedCrossRefGoogle Scholar
  19. Lueders T, Manefield M, Friedrich MW (2004) Enhanced sensitivity of DNA- and rRNA-based stable isotope probing by fractionation and quantitative analysis of isopycnic centrifugation gradients. Environ Microbiol 6:73–78PubMedCrossRefGoogle Scholar
  20. Luo G, Karakashev D, Xie L, Zhou Q, Angeridaki I (2011) Long-term effect of inoculum pretreatment on fermentative hydrogen production by repeated batch cultivations: homoacetogenesis and methanogenesis as competitors to hydrogen production. Biotechnol Bioeng 108:1816–1827PubMedCrossRefGoogle Scholar
  21. Nanqi R, Wanqian G, Bingfeng L, Guangli C, Jie D (2011) Biological hydrogen production by dark fermentation: challenges and prospects towards scaled-up production. Curr Opin Biotechnol 22:365–370CrossRefGoogle Scholar
  22. Patel MA, Ou MS, Harbrucker R, Aldrich HC, Buszko ML, Ingram LO, Shanmugam KT (2006) Isolation and characterization of acid-tolerant, thermophilic bacteria for effective fermentation of biomass-derived sugars to lactic acid. Appl Environ Microbiol 72:3228–3235PubMedCrossRefGoogle Scholar
  23. Rouvière P, Mandelco L, Winker S, Woese CR (1992) A detailed phylogeny for the Methanomicrobiales. Syst Appl Microbiol 15:363–371PubMedCrossRefGoogle Scholar
  24. Rozendal RA, Jeremiasse AW, Hamelers HVM, Buisman CJN (2008) Hydrogen production with a microbial biocathode. Environ Sci Technol 42:629–634PubMedCrossRefGoogle Scholar
  25. Sasaki K, Morita M, Hirano S, Ohmura N, Igarashi Y (2009) Effect of adding carbon fiber textiles to methanogenic bioreactors used to treat an artificial garbage slurry. J Biosci Bioeng 108:130–135PubMedCrossRefGoogle Scholar
  26. Sasaki K, Sasaki D, Morita M, Hirano S, Matsumoto N, Ohmura N, Igarashi Y (2010) Bioelectrochemical system stabilizes methane fermentation from garbage slurry. Bioresour Technol 101:3415–3422PubMedCrossRefGoogle Scholar
  27. Sasaki D, Hori T, Haruta S, Ueno Y, Ishii M, Igarashi Y (2011) Methanogenic pathway and community structure in a thermophilic anaerobic digestion process of organic solid waste. J Biosci Bioeng 111:41–46PubMedCrossRefGoogle Scholar
  28. Sasaki K, Morita M, Matsumoto N, Sasaki D, Hirano S, Ohmura N, Igarashi Y (2012a) Construction of hydrogen fermentation from garbage slurry using the membrane free bioelectrochemical system. J Biosci Bioeng 114:64–69CrossRefGoogle Scholar
  29. Sasaki K, Morita M, Sasaki D, Matsumoto N, Ohmura N, Igarashi Y (2012b) Single-chamber bioelectrochemical hydrogen fermentation from garbage slurry. Biochem Eng J 68:104–108CrossRefGoogle Scholar
  30. Sawayama S, Tsukahara K, Yagishita T (2006) Phylogenetic description of immobilized methanogenic community using real-time PCR in a fixed-bed anaerobic digester. Bioresour Technol 97:69–76PubMedCrossRefGoogle Scholar
  31. Soh ALA, Ralambotiana H, Olliver B, Prensier G, Tina E, Garcia JL (1991) Clostridium thermopalmarium sp. nov., a moderately thermophilic butyrate-producing bacterium isolated from palm wine in Senegal. Syst Appl Microbiol 14:135–139CrossRefGoogle Scholar
  32. Takai K, Horikoshi K (2000) Rapid detection and quantification of members of the archaeal community by quantitative PCR using fluorogenic probes. Appl Environ Microbiol 66:5066–5072PubMedCrossRefGoogle Scholar
  33. Thrash JG, Coates JD (2008) Review: Direct and indirect electrical stimulation of microbial metabolism. Environ Sci Technol 42:3921–3931PubMedCrossRefGoogle Scholar
  34. Ueno Y, Sasaki D, Fukui H, Haruta S, Ishii M, Igarashi Y (2006) Changes in bacterial community during fermentative hydrogen and acid production from organic waste by thermophilic anaerobic microflora. J Appl Microbiol 101:331–343PubMedCrossRefGoogle Scholar
  35. Ueno Y, Tatara M, Fukui H, Makiuchi T, Goto M, Sode K (2007) Production of hydrogen and methane from organic solid wastes by phase-separation of anaerobic process. Bioresour Technol 98:1861–1865PubMedCrossRefGoogle Scholar
  36. Updegraff DM (1969) Semimicro determination of cellulose in biological materials. Anal Biochem 32:420–424PubMedCrossRefGoogle Scholar
  37. Zeikus JG (1977) The biology of methanogenic bacteria. Bacteriol Rev 41:514–541PubMedGoogle Scholar
  38. Zinder SH, Sower KR, Ferry JG (1985) Methanosarcina thermophila sp. nov., a thermophilic, acetotrophic, methane-producing bacterium. Int J Syst Bacteriol 35:522–523CrossRefGoogle Scholar

Copyright information

© Springer-Verlag Berlin Heidelberg 2012

Authors and Affiliations

  • Kengo Sasaki
    • 1
    • 2
    • 3
  • Masahiko Morita
    • 1
    Email author
  • Daisuke Sasaki
    • 1
  • Naoya Ohmura
    • 1
  • Yasuo Igarashi
    • 2
  1. 1.Biotechnology Sector, Environmental Science Research LaboratoryCentral Research Institute of Electric Power IndustryAbiko-shiJapan
  2. 2.Department of Biotechnology, Graduate School of Agricultural and Life SciencesThe University of TokyoBunkyo-kuJapan
  3. 3.Organization of Advanced Science and TechnologyKobe UniversityKobeJapan

Personalised recommendations