Applied Microbiology and Biotechnology

, Volume 88, Issue 5, pp 1161–1167 | Cite as

Production of hexanoic acid from d-galactitol by a newly isolated Clostridium sp. BS-1

  • Byoung Seung Jeon
  • Byung-Chun Kim
  • Youngsoon Um
  • Byoung-In Sang
Applied Microbial and Cell Physiology


In a study screening anaerobic microbes utilizing d-galactitol as a fermentable carbon source, four bacterial strains were isolated from an enrichment culture producing H2, ethanol, butanol, acetic acid, butyric acid, and hexanoic acid. Among these isolates, strain BS-1 produced hexanoic acid as a major metabolic product of anaerobic fermentation with d-galactitol. Strain BS-1 belonged to the genus Clostridium based on phylogenetic analysis using 16S rRNA gene sequences, and the most closely related strain was Clostridium sporosphaeroides DSM 1294T, with 94.4% 16S rRNA gene similarity. In batch cultures, Clostridium sp. BS-1 produced 550 ± 31 mL L−1 of H2, 0.36 ± 0.01 g L−1 of acetic acid, 0.44 ± 0.01 g L−1 of butyric acid, and 0.98 ± 0.03 g L−1 of hexanoic acid in a 4-day cultivation. The production of hexanoic acid increased to 1.22 and 1.73 g L−1 with the addition of 1.5 g L−1 of sodium acetate and 100 mM 2-(N-morpholino)ethanesulfonic acid (MES), respectively. Especially when 1.5 g L−1 of sodium acetate and 100 mM MES were added simultaneously, the production of hexanoic acid increased up to 2.99 g L−1. Without adding sodium acetate, 2.75 g L−1 of hexanoic acid production from d-galactitol was achieved using a coculture of Clostridium sp. BS-1 and one of the isolates, Clostridium sp. BS-7, in the presence of 100 mM MES. In addition, volatile fatty acid (VFA) production by Clostridium sp. BS-1 from d-galactitol and d-glucose was enhanced when a more reduced culture redox potential (CRP) was applied via addition of Na2S·9H2O.


Clostridium sp. BS-1 Sludge Hexanoic acid d-galactitol Coculture 



This work was supported by the New & Renewable Energy of the Korea Institute of Energy Technology Evaluation and Planning (KETEP) grant funded by the Korea government Ministry of Knowledge Economy (No. 2009T100100337).

Supplementary material

253_2010_2827_MOESM1_ESM.pdf (1.2 mb)
Fig. S1 Total ion current chromatogram of VFAs produced in sludge culture broth. Peak no. 3 in the hexane extraction (A) and no 4. in the ethyl acetate extraction (B) are defined as hexanoic acid in a sludge culture utilizing d-galactitol. (PDF 1231 kb)


  1. Barker HA, Taha SM (1942) Clostridium kluyveri, an organism concerned in the formation of caproic acid from ethyl alcohol. J Bacteriol 43:347–363Google Scholar
  2. Collins MD, Lawson PA, Willems A, Cordoba JJ, Fernandez-Garayzabal J, Garcia P, Cai J, Hippe H, Farrow JA (1994) The phylogeny of the genus Clostridium: proposal of five new genera and eleven new species combinations. Int J Syst Bacteriol 44:812–826CrossRefGoogle Scholar
  3. Dürre P (2005) Handbook on clostridia, Taylor & FrancisGoogle Scholar
  4. Felsenstein J (1985) Confidence limits on phylogenies: an approach using the bootstrap. Evolution 39:783–791CrossRefGoogle Scholar
  5. Genthner BR, Davis CL, Bryant MP (1981) Features of rumen and sewage sludge strains of Eubacterium limosum, a methanol- and H2-CO2-utilizing species. Appl Environ Microbiol 42:12–19Google Scholar
  6. Herrero AA (1983) End-product inhibition in anaerobic fermentations. Trends Biotechnol 1:49–53CrossRefGoogle Scholar
  7. Holdeman LV, Cato EP, Moore WEC (1977) Anaerobe laboratory manual, 4th edn. Virginia Polytechnic Institute and State University, BlacksburgGoogle Scholar
  8. Hungate RE (1950) The anaerobic mesophilic cellulolytic bacteria. Bacteriol Rev 14:1–49Google Scholar
  9. Jeon B-S, Um Y, Lee S-M, Lee S-Y, Kim H-J, Kim YH, Gu MB, Sang B-I (2008) Performance analysis of a proton exchange membrane fuel cell (PEMFC) integrated with a trickling bed bioreactor for biological high-rate hydrogen production. Energy Fuels 22:83–86CrossRefGoogle Scholar
  10. Kawasaki S, Watamura Y, Ono M, Watanabe T, Takeda K, Niimura Y (2005) Adaptive responses to oxygen stress in obligatory anaerobes Clostridium acetobutylicum and Clostridium aminovalericum. Appl Environ Microbiol 71:8442–8450CrossRefGoogle Scholar
  11. Kenealy WR, Cao Y, Weimer PJ (1995) Production of caproic acid by cocultures of ruminal cellulolytic bacteria and Clostridium kluyveri grown on cellulose and ethanol. Appl Microbiol Biotechnol 44:507–513CrossRefGoogle Scholar
  12. Kim BH, Gadd GM (2008) Bacterial physiology and metabolism. Cambridge University Press, CambridgeCrossRefGoogle Scholar
  13. Kim BH, Bellows P, Datta R, Zeikus JG (1984) Control of carbon and electron flow in Clostridium acetobutylicum fermentations: utilization of carbon monoxide to inhibit hydrogen production and to enhance butanol yields. Appl Environ Microbiol 48:764–770Google Scholar
  14. Kleinert M, Barth T (2008) Towards a lignincellulosic biorefinery: direct one-step conversion of lignin to hydrogen-enriched biofuel. Energy Fuels 22:1371–1379CrossRefGoogle Scholar
  15. Kohlmiller EF Jr, Gest H (1951) A comparative study of the light and dark fermentations of organic acids by Rhodospirillum rubrum. J Bacteriol 61:269–282Google Scholar
  16. Kumar S, Tamura K, Nei M (2004) MEGA3: Integrated software for molecular evolutionary genetics analysis and sequence alignment. Brief Bioinform 5:150–163CrossRefGoogle Scholar
  17. 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
  18. Lee S-M, Cho MO, Park CH, Chung Y-C, Kim JH, Sang B-I, Um Y (2008) Continuous butanol production using suspended and immobilized Clostridium beijerinckii NCIMB 8052 with supplementary butyrate. Energy Fuels 22:3459–3464CrossRefGoogle Scholar
  19. Marounek M, Fliegrova K, Bartos S (1989) Metabolism and some characteristics of ruminal strains of Megasphaera elsdenii. Appl Environ Microbiol 55:1570–1573Google Scholar
  20. Nath K, Das D (2004) Improvement of fermentative hydrogen production: various approaches. Appl Microbiol Biotechnol 65:520–529CrossRefGoogle Scholar
  21. Ragauskas AJ, Williams CK, Davison BH et al (2006) The path forward for biofuels and biomaterials. Science 311:484–489CrossRefGoogle Scholar
  22. Robinson JW, Frame EMS, Frame GM (2004) Undergraduate instrumental analysis. Marcel Dekker, New YorkGoogle Scholar
  23. Roddick FA, Britz ML (1997) Production of hexanoic acid by free and immobilised cells of Megasphaera elsdenii: influence of in-situ product removal using Ion exchange resin. J Chem Technol Biotechnol 69:383–391CrossRefGoogle Scholar
  24. Saitou N, Nei M (1987) The neighbor-joining method: a new method for reconstructing phylogenetic trees. Mol Biol Evol 4:406–425Google Scholar
  25. Smith MV, Pierson MD (1979) Effect of reducing agents on oxidation-reduction potential and the outgrowth of Clostridium botulinum type E spores. Appl Environ Microbiol 37:978–984Google Scholar
  26. Thauer RK, Jungermann K, Henninger H, Wenning J, Decker H (1968) The energy metabolism of Clostridium kluyveri. Eur J Biochem 4:173–180CrossRefGoogle Scholar
  27. Thompson JD, Gibson TJ, Plewniak F, Jeanmougin F, Higgins DG (1997) The CLUSTAL_X windows interface: flexible strategies for multiple sequence alignment aided by quality analysis tools. Nucleic Acids Res 25:4876–4882CrossRefGoogle Scholar
  28. Wi SG, Kim HJ, Mahadevan SA, Yang DJ, Bae HJ (2009) The potential value of the seaweed Ceylon moss (Gelidium amansii) as an alternative bioenergy resource. Bioresour Technol 100:6658–6660CrossRefGoogle Scholar
  29. Wu Z, Yang S-T (2003) Extractive fermentation for butyric acid proruction from glucose by Clostridium tyrobutyricum. Biotechnol Bioeng 82:93–103CrossRefGoogle Scholar
  30. Zigova J, Sturdik E (2000) Advances in biotechnological production of butyric acid. J Ind Microbiol Biotechnol 24:153–160CrossRefGoogle Scholar

Copyright information

© Springer-Verlag 2010

Authors and Affiliations

  • Byoung Seung Jeon
    • 1
  • Byung-Chun Kim
    • 1
  • Youngsoon Um
    • 1
  • Byoung-In Sang
    • 1
  1. 1.Clean Energy CenterKorea Institute of Science and TechnologySeoulRepublic of Korea

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