Bioprocess and Biosystems Engineering

, Volume 42, Issue 2, pp 297–304 | Cite as

Improved cellulosic ethanol production from corn stover with a low cellulase input using a β-glucosidase-producing yeast following a dry biorefining process

  • Mesfin Geberekidan
  • Jian ZhangEmail author
  • Z. Lewis LiuEmail author
  • Jie Bao
Research Paper


A low-cost and sustainable cellulosic ethanol production is vital for fermentation-based industrial applications. Reducing the expenses of cellulose-deconstruction enzymes is one of the significant challenges to economic cellulose-to-ethanol conversion. Here, we report the improved ethanol production from corn stover after dry biorefining using a natural β-glucosidase-producing strain Clavispora NRRL Y-50464 with a low cellulase dose of 5 mg protein/g glucan under separate enzymatic hydrolysis and fermentation (SHF) and simultaneous saccharification and fermentation (SSF) conditions. Strain Clavispora NRRL Y-50464 exhibited a superior ethanol fermentation performance over Saccharomyces cerevisiae DQ1 under both conditions. It produced an ethanol titer of 38.1 g/L within 96 h at a conversion efficiency of 55.5% with 25% solids loading (w/w) via SSF without addition of extra β-glucosidase supplement. Improved performance of Y-50464 on a bioreactor with a helical stirring apparatus confirmed its advantage over the conventional bioreactors originally designed for liquid fermentations in cellulosic ethanol conversion by SSF. The results of this study suggested that the strain Clavispora NRRL Y-50464 has a potential as a candidate for lower-cost cellulosic ethanol production from lignocellulosic materials.


β-glucosidase Cellulosic ethanol Clavispora NRRL Y-50464 Corn stover Simultaneous saccharification and fermentation (SSF) 



This research was supported by the Natural Science Foundation of China (21306048) and sponsored by Shanghai Pujiang Program. Mention of trade names or commercial products in this publication is solely for the purpose of providing specific information and does not imply recommendation or endorsement by the US Department of Agriculture. USDA is an equal opportunity provider and employer.

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflict of interest.


  1. 1.
    Lynd L, Liang X, Biddy M, Allee A, Cai H, Foust T, Himmel M, Laser M, Wang M, Wyman C (2017) Cellulosic ethanol: status and innovation. Curr Opin Biotechnol 45:202–211Google Scholar
  2. 2.
    Zhang Q, Bao J (2017) Industrial cellulase performance in the simultaneous saccharification and co-fermentation (SSCF) of corn stover for high-titer ethanol production. Bioresour Bioprocess 4:17Google Scholar
  3. 3.
    Liu G, Zhang J, Bao J (2016) Cost evaluation of cellulase enzyme for industrial-scale cellulosic ethanol production based on rigorous Aspen Plus modeling. Bioprocess Biosyst Eng 39:133–140Google Scholar
  4. 4.
    Valdivia M, Galan J, Laffarga J, Ramos J (2016) Biofuels 2020: biorefineries based on lignocellulosic materials. Microb Biotechnol 9:585–594Google Scholar
  5. 5.
    Chauve M, Mathis H, Huc D, Casanave D, Monot F, Ferreira N (2010) Comparative kinetic analysis of two fungal β-glucosidases. Biotechnol Biofuels 3:3Google Scholar
  6. 6.
    Martins L, Kolling D, Camassola M, Dillon A, Ramos L (2008) Comparison of Penicillium echinulatum and Trichoderma reesei cellulases in relation to their activity against various cellulosic substrates. Bioresour Technol 99:1417–1424Google Scholar
  7. 7.
    Lee W, Nan H, Kim H, Jin Y (2013) Simultaneous saccharification and fermentation by engineered Saccharomyces cerevisiae without supplementing extracellular β-glucosidase. J Biotechnol 167:316–322Google Scholar
  8. 8.
    Haan R, Rensburg E, Rose S, Gorgens J, van Zyl W (2015) Progress and challenges in the engineering of non-cellulolytic microorganisms for consolidated bioprocessing. Curr Opin Biotechnol 33:32–38Google Scholar
  9. 9.
    Wang X, Liu ZL, Weber SA, Zhang X (2016) Two new native β-glucosidases from Clavispora NRRL Y-50464 confer its dual function as cellobiose fermenting ethanologenic yeast. PLoS One 11:e0151293Google Scholar
  10. 10.
    Liu ZL, Weber SA, Cotta M, Li S (2012) A new β-glucosidase producing yeast for lower-cost cellulosic ethanol production from xylose-extracted corncob residues by simultaneous saccharification and fermentation. Bioresour Technol 104:410–416Google Scholar
  11. 11.
    Liu ZL, Cotta M (2015) Technical assessment of cellulosic ethanol production using β-glucosidase producing yeast Clavispora NRRL Y-50464. Bioenerg Res 8:1203–1211Google Scholar
  12. 12.
    Zhang J, Chu D, Huang J, Yu Z, Dai G, Bao J (2010) Simultaneous saccharification and ethanol fermentation at high corn stover solids loading in a helical stirring bioreactor. Biotechnol Bioeng 105:718–728Google Scholar
  13. 13.
    Adney B, Baker J (1996) Measurement of cellulase activities. LAP-006. NREL analytical procedure. National Renewable Energy Laboratory, Golden COGoogle Scholar
  14. 14.
    Ghose T (1987) Measurement of cellulase activities. Pure Appl Chem 59:257–268Google Scholar
  15. 15.
    Bradford M (1976) A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem 25:248–256Google Scholar
  16. 16.
    Chu D, Zhang J, Bao J (2012) Simultaneous saccharification and ethanol fermentation of corn stover at high temperature and high solids loading by a thermotolerant strain Saccharomyces cerevisiae DQ1. Bioenerg Res 5:1020–1026Google Scholar
  17. 17.
    Zhang J, Wang X, Chu D, He Y, Bao J (2011) Dry pretreatment of lignocellulose with extremely low steam and water usage for bioethanol production. Bioresour Technol 102:4480–4488Google Scholar
  18. 18.
    He Y, Zhang L, Zhang J, Bao J (2014) Helically agitated mixing in dry dilute acid pretreatment enhances the bioconversion of corn stover into ethanol. Biotechnol Biofuels 7:1Google Scholar
  19. 19.
    Sluiter A, Hames B, Ruiz R, Scarlat C, Sluiter J, Templeton D, Crocker D (2008) Determination of structural carbohydrates and lignin in biomass. Technical Report NREL/TP-510-42618. Laboratory Analytical Procedure (LAP), Golden COGoogle Scholar
  20. 20.
    Zhang J, Zhu Z, Wang N, Wang W, Bao J (2010) Biodetoxification of toxins generated from lignocellulose pretreatment using a newly isolated fungus Amorphotheca resinae ZN1 and the consequent ethanol fermentation. Biotechnol Biofuels 3:26Google Scholar
  21. 21.
    He Y, Zhang J, Bao J (2016) Acceleration of biodetoxification on dilute acid pretreated lignocellulose feedstock by aeration and the consequent ethanol fermentation evaluation. Biotechnol Biofuels 9:19Google Scholar
  22. 22.
    Zhang J, Lei C, Liu G, Bao Y, Balan V, Bao J (2017) In-situ vacuum distillation of ethanol helps to recycle cellulase and yeast during SSF of delignified corncob residues. ACS Sustain Chem Eng 5:11676–11685Google Scholar
  23. 23.
    Zhang J, Bao J (2012) A modified method for calculating practical ethanol yield at high lignocellulosic solids content and high ethanol titer. Bioresour Technol 116:74–79Google Scholar
  24. 24.
    Olofsson K, Bertilsson M, Liden G (2008) A short review on SSF-an interesting process option for ethanol production from lignocellulosic feedstocks. Biotechnol Biofuels 1:7Google Scholar
  25. 25.
    Liu ZL, Weber SA, Cotta M (2013) Isolation and characterization of a β-glucosidase from a Clavispora strain with potential applications in bioethanol production from cellulosic materials. Bioenerg Res 6:65–74Google Scholar
  26. 26.
    Shen Y, Zhang Y, Ma T, Bao X, Du F, Zhuang G, Qu Y (2008) Simultaneous saccharification and fermentation of acid-pretreated corncobs with a recombinant Saccharomyces cerevisiae expressing β-glucosidase. Bioresour Technol 99:5099–5103Google Scholar
  27. 27.
    Treebupachatsakul T, Nakazawa H, Shinbo H, Fujikawa H, Nagaiwa A, Ochiai N, Kawaguchi T, Nikaido M, Totani K, Shioya K, Shida Y, Morikawa Y, Ogasawara W, Okada H (2016) Heterologously expressed Aspergillus aculeatus β-glucosidase in Saccharomyces cerevisiae is a cost-effective alternative to commercial supplementation of β-glucosidase in industrial ethanol production using Trichoderma reesei cellulases. J Biosci Bioeng 121:27–35Google Scholar
  28. 28.
    Hu M, Zha J, He L, Lv Y, Shen M, Zhong C, Li B, Yuan Y (2016) Enhanced bioconversion of cellobiose by industrial Saccharomyces cerevisiae used for cellulose utilization. Front Microbiol 7:241Google Scholar
  29. 29.
    Chapla D, Parikh B, Liu L, Cotta M, Kumar A (2015) Enhanced cellulosic ethanol production from mild-alkali pretreated rice straw in SSF using Clavispora NRRL Y-50464. J Biobased Mater Bioenergy 9:1–8Google Scholar
  30. 30.
    Kumar AK, Parikh BS, Shah E, Liu ZL, Cotta MA (2016) Cellulosic ethanol production from green solvent-pretreated rice straw. Biocatal Agric Biotechnol 7:14–23Google Scholar

Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2018

Authors and Affiliations

  1. 1.State Key Laboratory of Bioreactor EngineeringEast China University of Science and TechnologyShanghaiChina
  2. 2.Bioenergy Research UnitNational Center for Agricultural Utilization Research, USDA-ARSPeoriaUSA

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