Production of optically pure 2,3-butanediol from Miscanthus floridulus hydrolysate using engineered Bacillus licheniformis strains

  • Yabin Gao
  • Huahua Huang
  • Shouwen Chen
  • Gaofu QiEmail author
Original Paper


2,3-Butanediol (2,3-BD) can be produced by fermentation of natural resources like Miscanthus. Bacillus licheniformis mutants, WX-02ΔbudC and WX-02ΔgldA, were elucidated for the potential to use Miscanthus as a cost-effective biomass to produce optically pure 2,3-BD. Both WX-02ΔbudC and WX-02ΔgldA could efficiently use xylose as well as mixed sugars of glucose and xylose to produce optically pure 2,3-BD. Batch fermentation of M. floridulus hydrolysate could produce 21.6 g/L d-2,3-BD and 23.9 g/L meso-2,3-BD in flask, and 13.8 g/L d-2,3-BD and 13.2 g/L meso-2,3-BD in bioreactor for WX-02ΔbudC and WX-02ΔgldA, respectively. Further fed-batch fermentation of hydrolysate in bioreactor showed both of two strains could produce optically pure 2,3-BD, with 32.2 g/L d-2,3-BD for WX-02ΔbudC and 48.5 g/L meso-2,3-BD for WX-02ΔgldA, respectively. Collectively, WX-02ΔbudC and WX-02ΔgldA can efficiently produce optically pure 2,3-BD with M. floridulus hydrolysate, and these two strains are candidates for industrial production of optical purity of 2,3-BD with M. floridulus hydrolysate.


Bacillus licheniformis Optical purity d-2,3-Butanediol meso-2,3-Butanediol Miscanthus floridulus 


  1. Biswas R, Yamaoka M, Nakayama H, Kondo T, Yoshida K, Bisaria VS, Kondo A (2012) Enhanced production of 2,3-butanediol by engineered Bacillus subtilis. Appl Microbiol Biotechnol 94, 651–658CrossRefPubMedGoogle Scholar
  2. Chen T, Liu WX, Fu J, Zhang B, Tang YJ (2013) Engineering Bacillus subtilis for acetoin production from glucose and xylose mixtures. J Biotechnol 168:499–505CrossRefPubMedGoogle Scholar
  3. Chirino-Valle I, Kandula D, Littlejohn C, Hill R, Walker M, Shields M, Cummings N, Hettiarachchi D, Wratten S (2016) Potential of the beneficial fungus Trichoderma to enhance ecosystem-service provision in the biofuel grass Miscanthus × giganteus in agriculture. Sci Rep 6:25109CrossRefPubMedPubMedCentralGoogle Scholar
  4. Chu H, Xin B, Liu P, Wang Y, Li L, Liu X, Zhang X, Ma C, Ping Xu, Gao C (2015) Metabolic engineering of Escherichia coli for production of (2S, 3S)-2,3-butanediol from glucose. Biotechnol Biofuels 8:143–154CrossRefPubMedPubMedCentralGoogle Scholar
  5. Fu J, Wang Z, Chen T, Liu W, Shi T, Wang G, Tang YJ, Zhao X (2014) NADH plays the vital role for chiral pure d-2,3-butanediol production in Bacillus subtilis under limited oxygen conditions. Biotechnol Bioeng 111:2126–2131CrossRefPubMedGoogle Scholar
  6. Fu J, Hu G, Feng L, Mao Y, Wang Z, Ma H, Chen T, Zhao X (2016) Metabolic engineering of Bacillus subtilis for chiral pure meso-2,3-butanediol production. Biotechnol Biofuels 9:90–104CrossRefPubMedPubMedCentralGoogle Scholar
  7. Huang CF, Jiang YF, Guo GL, Hwang WS (2013) Method of 2,3-butanediol production from glycerol and acid-pretreated rice straw hydrolysate by newly isolated strains: pre-evaluation as an integrated biorefinery process. Bioresour Technol 135:446–453CrossRefPubMedGoogle Scholar
  8. Ji XJ, Liu LG, Shen MQ, Nie ZK, Tong YJ, Huang H (2015) Constructing a synthetic metabolic pathway in Escherichia coli to produce the enantiomerically pure (R,R)-2,3-butanediol. Biotechnol Bioeng 112:1056–1059CrossRefPubMedGoogle Scholar
  9. Jiang LQ, Fang Z, Guo F, Yang LB (2012) Production of 2,3-butanediol from acid hydrolysates of Jatropha hulls with Klebsiella oxytoca. Bioresour Technol 107:405–410CrossRefPubMedGoogle Scholar
  10. Jurchescu IM, Hamann J, Zhou X, Ortmann T, Kuenz A, Prüße U, Lang S (2013) Enhanced 2,3-butanediol production in fed-batch cultures of free and immobilized Bacillus licheniformis DSM 8785. Appl Microbiol Biotechnol 97:6715–6723CrossRefPubMedGoogle Scholar
  11. Lee WC, Kuan WC (2015) Miscanthus as cellulosic biomass for bioethanol production. Biotechnol J 10:840–854CrossRefPubMedGoogle Scholar
  12. Lee S, Kim B, Park K, Um Y, Lee J (2012) Synthesis of pure meso-2,3-butanediol from crude glycerol using an engineered metabolic pathway in Escherichia coli. Appl Biochem Biotechnol 166:1801–1813CrossRefPubMedGoogle Scholar
  13. Lewandowska M, Szymańska K, Kordala N, Dąbrowska A, Bednarski W, Juszczuk A (2016) Evaluation of Mucor indicus and Saccharomyces cerevisiae capability to ferment hydrolysates of rape straw and Miscanthus giganteus as affected by the pretreatment method. Bioresour Technol 212:262 – 70CrossRefPubMedGoogle Scholar
  14. Li L, Wang Y, Zhang L, Ma C, Wang A, Tao F, Xu P (2012) Biocatalytic production of (2S,3S)-2,3-butanediol from diacetyl using whole cells of engineered Escherichia coli. Bioresour Technol 115:111–116CrossRefPubMedGoogle Scholar
  15. Li J, Wang W, Ma Y, Zeng AP (2013a) Medium optimization and proteome analysis of (R,R)-2,3-butanediol production by Paenibacillus polymyxa ATCC 12321. Appl Microbiol Biotechnol 97:585–597CrossRefPubMedGoogle Scholar
  16. Li L, Zhang L, Li K, Wang Y, Gao C, Han B, Ma C, Xu P (2013b) A newly isolated Bacillus licheniformis strain thermophilically produces 2,3-butanediol, a platform and fuel bio-chemical. Biotechnol Biofuels 6:123CrossRefPubMedPubMedCentralGoogle Scholar
  17. Li L, Li K, Wang K, Chen C, Gao C, Ma C, Xu P (2014) Efficient production of 2,3-butanediol from corn stover hydrolysate by using a thermophilic Bacillus licheniformis strain. Bioresour Technol 170:256–261CrossRefPubMedGoogle Scholar
  18. Liu Z, Qin J, Gao C, Hua D, Ma C, Li L, Wang Y, Xu P (2011) Production of (2S,3S)-2,3-butanediol and (3S)-acetoin from glucose using resting cells of Klebsiella pneumonia and Bacillus subtilis. Bioresour Technol 102:10741–10744CrossRefPubMedGoogle Scholar
  19. Mazumdar S, Lee J, Oh MK (2013) Microbial production of 2,3 butanediol from seaweed hydrolysate using metabolically engineered Escherichia coli. Bioresour Technol 136:329–336CrossRefPubMedGoogle Scholar
  20. Qi G, Kang Y, Li L, Xiao A, Zhang S, Wen Z, Xu D, Chen S (2014) Deletion of meso-2,3-butanediol dehydrogenase gene budC for enhanced d-2,3-butanediol production in Bacillus licheniformis. Biotechnol Biofuels 7:16CrossRefPubMedPubMedCentralGoogle Scholar
  21. Qiu Y, Zhang J, Li L, Wen Z, Nomura CT, Wu S, Chen S (2016) Engineering Bacillus licheniformis for the production of meso-2,3-butanediol. Biotechnol Biofuels 9:117CrossRefPubMedPubMedCentralGoogle Scholar
  22. Roberts SB, Gowen CM, Brooks JP, Fong SS (2010) Genome-scale metabolic analysis of Clostridium thermocellum for bioethanol production. BMC systems Biology 4:31CrossRefPubMedPubMedCentralGoogle Scholar
  23. Siemerink MAJ, Kuit W, Contreras AML, van der Oost EGJ, Kenge SWM (2011) d-2,3-butanediol production due to heterologous expression of an acetoin reductase in Clostridium acetobutylicum. Appl Environ Microbiol 77:2582–2588CrossRefPubMedPubMedCentralGoogle Scholar
  24. Świątek K, Lewandowska M, Świątek M, Bednarski W, Brzozowski B (2014) The improvement of enzymatic hydrolysis efficiency of rape straw and Miscanthus giganteus polysaccharides. Bioresour Technol 151:323–331CrossRefPubMedGoogle Scholar
  25. Tong YJ, Ji XJ, Shen MQ, Liu LG, Nie ZK, Huang H (2016) Constructing a synthetic constitutive metabolic pathway in Escherichia coli for (R, R)-2,3-butanediol production. Appl Microbiol Biotechnol 100:637 – 47CrossRefPubMedGoogle Scholar
  26. Wang A, Wang Y, Jiang T, Li L, Ma C, Xu P (2010) Production of 2,3-butanediol from corncob molasses, a waste by-product in xylitol production. Appl Microbiol Biotechnol 87:965–970CrossRefPubMedGoogle Scholar
  27. Wang Q, Chen T, Zhao X, Chamu J (2012) Metabolic engineering of thermophilic Bacillus licheniformis for chiral pure d-2,3-butanediol production. Biotechnol Bioeng 109:1610–1621CrossRefPubMedGoogle Scholar
  28. Wu G, Yan Q, Jones JA, Tang YJ, Fong SS, Koffas MAG (2016) Metabolic burden: cornerstones in synthetic biology and metabolic engineering applications. Trends Biotechnol 34:652–664CrossRefPubMedGoogle Scholar
  29. Xu Y, Chu H, Gao C, Tao F, Zhou Z, Li K, Li L, Ma C, Xu P (2014) Systematic metabolic engineering of Escherichia coli for high-yield production of fuel bio-chemical 2,3-butanediol. Metab Eng 23:22–33CrossRefPubMedGoogle Scholar
  30. Yan Q, Hong E, Fong SS (2017) Study of ChiR function in Serratia marcescens and its application for improving 2,3-butanediol from crystal chitin. Appl Microbiol Biotechnol 101(19):1–12Google Scholar
  31. Yang T, Rao Z, Zhang X, Lin Q, Xia H, Xu Z, Yang S (2011a) Production of 2,3-butanediol from glucose by GRAS microorganism Bacillus amyloliquefaciens. J Basic Microbiol 51:650–658CrossRefPubMedGoogle Scholar
  32. Yang T, Zhang X, Rao Z, Gu S, Xia H, Xu Z (2011b) Optimization and scale-up of 2,3-butanediol production by Bacillus amyloliquefaciens B10-127. World J Microb Biotechnol 28:1563–1574CrossRefGoogle Scholar
  33. Yang T, Rao Z, Zhang X, Xu M, Xu Z, Yang ST (2013a) Improved production of 2,3-Butanediol in Bacillus amyloliquefaciens by over-expression of glyceraldehyde-3-phosphate dehydrogenase and 2,3-butanediol dehydrogenase. PLoS ONE 8:e76149CrossRefPubMedPubMedCentralGoogle Scholar
  34. Yang TW, Rao ZM, Zhang X, Xu MJ, Xu ZH, Yang ST (2013b) Effects of corn steep liquor on production of 2,3-butanediol and acetoin by Bacillus subtilis. Process Biochem 48:1610–1617CrossRefGoogle Scholar
  35. Yang TW, Rao ZM, Zhang X, Xu MJ, Xu ZH, Yang ST (2013c) Fermentation of biodiesel-derived glycerol by Bacillus amyloliquefaciens: effects of co-substrates on 2,3-butanediol production. Appl Microbiol Biotechnol 97:7651–7658CrossRefPubMedGoogle Scholar

Copyright information

© Springer Science+Business Media B.V., part of Springer Nature 2018

Authors and Affiliations

  • Yabin Gao
    • 1
  • Huahua Huang
    • 1
  • Shouwen Chen
    • 1
  • Gaofu Qi
    • 1
    Email author
  1. 1.College of Life Science and TechnologyHuazhong Agricultural UniversityWuhanPeople’s Republic of China

Personalised recommendations