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Fermentation of deproteinized cheese whey powder solutions to ethanol by engineered Saccharomyces cerevisiae: effect of supplementation with corn steep liquor and repeated-batch operation with biomass recycling by flocculation

  • Original Paper
  • Published:
Journal of Industrial Microbiology & Biotechnology

Abstract

The lactose in cheese whey is an interesting substrate for the production of bulk commodities such as bio-ethanol, due to the large amounts of whey surplus generated globally. In this work, we studied the performance of a recombinant Saccharomyces cerevisiae strain expressing the lactose permease and intracellular ß-galactosidase from Kluyveromyces lactis in fermentations of deproteinized concentrated cheese whey powder solutions. Supplementation with 10 g/l of corn steep liquor significantly enhanced whey fermentation, resulting in the production of 7.4% (v/v) ethanol from 150 g/l initial lactose in shake-flask fermentations, with a corresponding productivity of 1.2 g/l/h. The flocculation capacity of the yeast strain enabled stable operation of a repeated-batch process in a 5.5-l air-lift bioreactor, with simple biomass recycling by sedimentation of the yeast flocs. During five consecutive batches, the average ethanol productivity was 0.65 g/l/h and ethanol accumulated up to 8% (v/v) with lactose-to-ethanol conversion yields over 80% of theoretical. Yeast viability (>97%) and plasmid retention (>84%) remained high throughout the operation, demonstrating the stability and robustness of the strain. In addition, the easy and inexpensive recycle of the yeast biomass for repeated utilization makes this process economically attractive for industrial implementation.

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References

  1. Agarwal L, Dutt K, Meghwanshi GK, Saxena RK (2008) Anaerobic fermentative production of lactic acid using cheese whey and corn steep liquor. Biotechnol Lett 30:631–635

    Article  PubMed  CAS  Google Scholar 

  2. Amartey S, Jeffries TW (1994) Comparison of corn steep liquor with other nutrients in the fermentation of D-xylose by Pichia stipitis CBS 6054. Biotechnol Lett 16:211–214

    Article  CAS  Google Scholar 

  3. Casey GP, Ingledew WMM (1986) Ethanol tolerance in yeasts. CRC Crit Rev Microbiol 13:219–280

    Article  CAS  Google Scholar 

  4. Choi GW, Kang HW, Moon SK (2009) Repeated-batch fermentation using flocculent hybrid, Saccharomyces cerevisiae CHFY0321 for efficient production of bioethanol. Appl Microbiol Biotechnol 84:261–269

    Article  PubMed  CAS  Google Scholar 

  5. Compagno C, Porro D, Smeraldi C, Ranzi BM (1995) Fermentation of whey and starch by transformed Saccharomyces cerevisiae cells. Appl Microbiol Biotechnol 43:822–825

    Article  PubMed  CAS  Google Scholar 

  6. Domingues L, Guimarães PMR, Oliveira C (2010) Metabolic engineering of Saccharomyces cerevisiae for lactose/whey fermentation. Bioengineered Bugs 1 (in press)

  7. Domingues L, Lima N, Teixeira JA (2001) Alcohol production from cheese whey permeate using genetically modified flocculent yeast cells. Biotechnol Bioeng 72:507–514

    Article  PubMed  CAS  Google Scholar 

  8. Domingues L, Teixeira JA, Lima N (1999) Construction of a flocculent Saccharomyces cerevisiae fermenting lactose. Appl Microbiol Biotechnol 51:621–626

    Article  PubMed  CAS  Google Scholar 

  9. Gawel J, Kosikowski FV (1978) Improving alcohol fermentation in concentrated ultrafiltration permeates of cottage cheese whey. J Food Sci 43:1717–1719

    Article  CAS  Google Scholar 

  10. Ghaly AE, El-Taweel AA (1997) Continuous ethanol production from cheese whey fermentation by Candida pseudotropicalis. Energy Sources A Recover Util Environ 19:1043–1063

    Article  CAS  Google Scholar 

  11. Ghaly AE, El-Taweel AA (1995) Effect of micro-aeration on the growth of Candida pseudotropicalis and production of ethanol during batch fermentation of cheese whey. Bioresour Technol 52:203–217

    Article  CAS  Google Scholar 

  12. Grba S, Stehlik-Tomas V, Stanzer D, Vahcic N, Skrlin A (2002) Selection of yeast strain Kluyveromyces marxianus for alcohol and biomass production on whey. Chem Biochem Eng Q 16:13–16

    CAS  Google Scholar 

  13. Guimarães PMR, François J, Parrou JL, Teixeira JA, Domingues L (2008) Adaptive evolution of a lactose-consuming Saccharomyces cerevisiae recombinant. Appl Environ Microbiol 74:1748–1756

    Article  PubMed  CAS  Google Scholar 

  14. Guimarães PMR, Teixeira JA, Domingues L (2008) Fermentation of high concentrations of lactose to ethanol by engineered flocculent Saccharomyces cerevisiae. Biotechnol Lett 30:1953–1958

    Article  PubMed  CAS  Google Scholar 

  15. Guimarães PMR, Teixeira JA, Domingues L (2010) Fermentation of lactose to bio-ethanol by yeasts as part of integrated solutions for the valorisation of cheese whey. Biotechnol Adv 28:375–384

    Article  PubMed  CAS  Google Scholar 

  16. Guimarães PMR, Virtanen H, Londesborough J (2006) Direct evidence that maltose transport activity is affected by the lipid composition of brewer’s yeast. J Inst Brew 112:203–209

    Google Scholar 

  17. Guo X, Zhou J, Xiao D (2010) Improved ethanol production by mixed immobilized cells of Kluyveromyces marxianus and Saccharomyces cerevisiae from cheese whey powder solution fermentation. Appl Biochem Biotechnol 160:532–538

    Article  PubMed  CAS  Google Scholar 

  18. Hahn-Hägerdal B, Karhumaa K, Larsson CU, Gorwa-Grauslund M, Gorgens J, van Zyl WH (2005) Role of cultivation media in the development of yeast strains for large-scale industrial use. Microb Cell Fact 4:31

    Article  PubMed  CAS  Google Scholar 

  19. Huang Y, Yang ST (1998) Acetate production from whey lactose using co-immobilized cells of homolactic and homoacetic bacteria in a fibrous-bed bioreactor. Biotechnol Bioeng 60:498–507

    Article  PubMed  CAS  Google Scholar 

  20. Janssens JH, Burris N, Woodward A, Bailey RB (1983) Lipid-enhanced ethanol production by Kluyveromyces fragilis. Appl Environ Microbiol 45:598–602

    PubMed  CAS  Google Scholar 

  21. Kadam KL, Newman MM (1997) Development of a low-cost fermentation medium for ethanol production from biomass. Appl Microbiol Biotechnol 47:625–629

    Article  PubMed  CAS  Google Scholar 

  22. Kargi F, Ozmihci S (2006) Utilization of cheese whey powder (CWP) for ethanol fermentations: effects of operating parameters. Enzyme Microbiol Technol 38:711–718

    Article  CAS  Google Scholar 

  23. Kim HO, Wee YJ, Kim JN, Yun JS, Ryu HW (2006) Production of lactic acid from cheese whey by batch and repeated batch cultures of Lactobacillus sp. RKY2. Appl Biochem Biotechnol 131:694–704

    Article  PubMed  Google Scholar 

  24. Klein J, Maia J, Vicente AA, Domingues L, Teixeira JA, Jurascik M (2005) Relationships between hydrodynamics and rheology of flocculating yeast suspensions in a high-cell-density airlift bioreactor. Biotechnol Bioeng 89:393–399

    Article  PubMed  CAS  Google Scholar 

  25. Koutinas AA, Athanasiadis I, Bekatorou A et al (2007) Kefir-yeast technology: industrial scale-up of alcoholic fermentation of whey, promoted by raisin extracts, using kefir-yeast granular biomass. Enzyme Microbiol Technol 41:576–582

    Article  CAS  Google Scholar 

  26. Lawford HG, Rousseau JD (1997) Corn steep liquor as a cost-effective nutrition adjunct in high-performance Zymomonas ethanol fermentations. Appl Biochem Biotechnol 63–5:287–304

    Article  Google Scholar 

  27. Lee PC, Lee SY, Hong SH, Chang HN (2003) Batch and continuous cultures of Mannheimia succiniciproducens MBEL55E for the production of succinic acid from whey and corn steep liquor. Bioprocess Biosyst Eng 26:63–67

    Article  PubMed  CAS  Google Scholar 

  28. Li F, Zhao XQ, Ge XM, Bai FW (2009) An innovative consecutive batch fermentation process for very high gravity ethanol fermentation with self-flocculating yeast. Appl Microbiol Biotechnol 84:1079–1086

    Article  PubMed  CAS  Google Scholar 

  29. Ling KC (2008) Whey to ethanol: a biofuel role for dairy cooperatives? USDA Rural Development, Research Report 214, Washington, DC

  30. Ma K, Wakisaka M, Sakai K, Shirai Y (2009) Flocculation characteristics of an isolated mutant flocculent Saccharomyces cerevisiae strain and its application for fuel ethanol production from kitchen refuse. Bioresour Technol 100:2289–2292

    Article  PubMed  CAS  Google Scholar 

  31. Mahmoud MM, Kosikowski FV (1982) Alcohol and single cell protein production by Kluyveromyces in concentrated whey permeates with reduced ash. J Dairy Sci 65:2082–2087

    Article  CAS  Google Scholar 

  32. Miller GL (1959) Use of dinitrosalicylic acid reagent for determination of reducing sugar. Anal Chem 31:426–428

    Article  CAS  Google Scholar 

  33. Mills DR (1941) Differential staining of living and dead yeast cells. Food Res 6:361–371

    Google Scholar 

  34. Oda Y, Nakamura K (2009) Production of ethanol from the mixture of beet molasses and cheese whey by a 2-deoxyglucose-resistant mutant of Kluyveromyces marxianus. FEMS Yeast Res 9:742–748

    Article  PubMed  CAS  Google Scholar 

  35. Ozmihci S, Kargi F (2007) Effects of feed sugar concentration on continuous ethanol fermentation of cheese whey powder solution (CWP). Enzyme Microbiol Technol 41:876–880

    Article  CAS  Google Scholar 

  36. Ozmihci S, Kargi F (2007) Ethanol fermentation of cheese whey powder solution by repeated fed-batch operation. Enzyme Microbiol Technol 41:169–174

    Article  CAS  Google Scholar 

  37. Ozmihci S, Kargi F (2007) Kinetics of batch ethanol fermentation of cheese-whey powder (CWP) solution as function of substrate and yeast concentrations. Bioresour Technol 98:2978–2984

    Article  PubMed  CAS  Google Scholar 

  38. Porro D, Martegani E, Ranzi BM, Alberghina L (1992) Lactose/whey utilization and ethanol production by transformed Saccharomyces cerevisiae cells. Biotechnol Bioeng 39:799–805

    Article  PubMed  CAS  Google Scholar 

  39. Ramakrishnan S, Hartley BS (1993) Fermentation of lactose by yeast cells secreting recombinant fungal lactase. Appl Environ Microbiol 59:4230–4235

    PubMed  CAS  Google Scholar 

  40. Rigoulet M, Aguilaniu H, Averet N et al (2004) Organization and regulation of the cytosolic NADH metabolism in the yeast Saccharomyces cerevisiae. Mol Cell Biochem 256–257:73–81

    Article  PubMed  Google Scholar 

  41. Rosenberg M, Tomaska M, Kanuch J, Sturdik E (1995) Improved ethanol production from whey with Saccharomyces cerevisiae using permeabilized cells of Kluyveromyces marxianus. Acta Biotechnol 15:387–390

    Article  CAS  Google Scholar 

  42. Roy D, Goulet J, Leduy A (1986) Batch fermentation of whey ultrafiltrate by Lactobacillus helveticus for lactic acid production. Appl Microbiol Biotechnol 24:206–213

    Article  CAS  Google Scholar 

  43. Silveira WB, Passos F, Mantovani HC, Passos FML (2005) Ethanol production from cheese whey permeate by Kluyveromyces marxianus UFV-3: a flux analysis of oxido-reductive metabolism as a function of lactose concentration and oxygen levels. Enzyme Microbiol Technol 36:930–936

    Article  CAS  Google Scholar 

  44. Siso MIG (1996) The biotechnological utilization of cheese whey: a review. Bioresour Technol 57:1–11

    Article  Google Scholar 

  45. Smithers GW (2008) Whey and whey proteins—from ‘gutter-to-gold’. Int Dairy J 18:695–704

    Article  CAS  Google Scholar 

  46. Szczodrak J, Szewczuk D, Rogalski J, Fiedurek J (1997) Selection of yeast strain and fermentation conditions for high-yield ethanol production from lactose and concentrated whey. Acta Biotechnologica 17:51–61

    Article  CAS  Google Scholar 

  47. Tang Y, An M, Liu K et al (2006) Ethanol production from acid hydrolysate of wood biomass using the flocculating yeast Saccharomyces cerevisiae strain KF-7. Process Biochem 41:909–914

    Article  CAS  Google Scholar 

  48. Verduyn C, Postma E, Scheffers WA, van Dijken JP (1992) Effect of benzoic acid on metabolic fluxes in yeasts: a continuous-culture study on the regulation of respiration and alcoholic fermentation. Yeast 8:501–517

    Article  PubMed  CAS  Google Scholar 

  49. Zhang J, Reddy J, Buckland B, Greasham R (2003) Toward consistent and productive complex media for industrial fermentations: studies on yeast extract for a recombinant yeast fermentation process. Biotechnol Bioeng 82:640–652

    Article  PubMed  CAS  Google Scholar 

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Acknowledgments

The authors thank André Mota for assistance in setting up the bioreactor fermentations, LACTOGAL—Produtos Alimentares S.A. (Portugal) for kindly providing the cheese whey powder and COPAM—Companhia Portuguesa de Amidos, S.A. (Portugal) for kindly providing the CSL. The financial support of Fundação para a Ciência e a Tecnologia (FCT), Portugal, is acknowledged (project ProBioethanol PTDC/BIO/66151/2006 and grant SFRH/BPD/44328/2008 to P. M. R. Guimarães).

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Authors and Affiliations

  1. IBB-Institute for Biotechnology and Bioengineering, Centre of Biological Engineering, Universidade do Minho, Campus de Gualtar, 4710-057, Braga, Portugal

    Ana Carina Silva, Pedro M. R. Guimarães, José A. Teixeira & Lucília Domingues

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  1. Ana Carina Silva
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  2. Pedro M. R. Guimarães
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  3. José A. Teixeira
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  4. Lucília Domingues
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Correspondence to Lucília Domingues.

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Silva, A.C., Guimarães, P.M.R., Teixeira, J.A. et al. Fermentation of deproteinized cheese whey powder solutions to ethanol by engineered Saccharomyces cerevisiae: effect of supplementation with corn steep liquor and repeated-batch operation with biomass recycling by flocculation. J Ind Microbiol Biotechnol 37, 973–982 (2010). https://doi.org/10.1007/s10295-010-0748-z

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