Applied Biochemistry and Biotechnology

, Volume 166, Issue 1, pp 36–46 | Cite as

Optimization of Laccase Production by Trametes versicolor Cultivated on Industrial Waste

  • Marina Tišma
  • Polona Žnidaršič-Plazl
  • Đurđa Vasić-Rački
  • Bruno ZelićEmail author


Laccases are very interesting biocatalysts for several industrial applications. Its production by different white-rot fungi can be stimulated by a variety of inducing substrates, and the use of lignocellulosic wastes or industrial by-products is one of the possible approaches to reduce production costs. In this work, various industrial wastes were tested for laccase production by Trametes versicolor MZKI G-99. Solid waste from chemomechanical treatment facility of a paper manufacturing plant showed the highest potential for laccase production. Enzyme production during submerged cultivation of T. versicolor on the chosen industrial waste has been further improved by medium optimization using genetic algorithm. Concentrations of five components in the medium were optimized within 60 shake-flasks experiments, where the highest laccase activity of 2,378 U dm−3 was achieved. Waste from the paper industry containing microparticles of CaCO3 was found to stimulate the formation of freely dispersed mycelium and laccase production during submerged cultivation of T. versicolor. It was proven to be a safe and inexpensive substrate for commercial production of laccase and might be more widely applicable for metabolite production by filamentous fungi.


Laccase production Industrial waste Medium optimization Genetic algorithm Trametes versicolor Microparticles 



This work was supported by the Croatian Ministry of Science, Education, and Sports (contract grant number 125-1252086-2793) and by The National Foundation for Science, Higher Education, and Technological Development of the Republic of Croatia (Program NZZ Installation Grant). P. Žnidaršič Plazl was supported by grant P2-0191, provided by the Ministry of Higher Education, Science, and Technology of the Republic of Slovenia. The authors gratefully acknowledges Dr. D. Ravnjak for the provision of CaCO3, sludge, and pulps from Papirnica Vevče, Ljubljana, Slovenia; Mrs. I. Škraba for providing microorganism from the Microbial Culture Collection of the National Institute of Chemistry, Slovenia; Mrs. Mira Špehar for the provision of the waste from the malt industry and barley husk, Slavonija slad d.o.o., Nova Gradiška, Croatia; and Mr. Dean Pinjuh for the provision of sawdusts, Hrvatske šume d.o.o., Slavonski Brod, Croatia. The authors wish to thank Mrs. Nataša Car for the skilled technical assistance.


  1. 1.
    Asgher, M., Bhatti, H. N., Ashraf, M., & Legge, R. L. (2008). Recent developments in biodegradation of industrial pollutants by white rot fungi and their enzyme system. Biodegradation, 19, 771–783.CrossRefGoogle Scholar
  2. 2.
    Schlosser, D., Grey, R., & Fritsche, W. (1997). Patents of lignolytic enzymes in Trametes versicolor. Distribution of extra- and intracellular enzyme activities during cultivation on glucose wheat straw and beech wood. Applied Microbiology and Biotechnology, 47, 412–418.CrossRefGoogle Scholar
  3. 3.
    Pozdnyakova, N., Leontievsky, A., & Golovleva, L. (1997). Oxidase of the white rot fungus Panus tigrinus. FEBS Letters, 350, 192–194.CrossRefGoogle Scholar
  4. 4.
    Teerapatsakul, C., Parra, R., Bucke, C., & Chitradon, L. (2007). Improvement of laccase production from Ganoderma sp. KU-Alk4 by medium engineering. World Journal of Microbiology and Biotechnology, 23, 1519–1527.CrossRefGoogle Scholar
  5. 5.
    Moreira, M. T., Feijoo, G., & Lema, J. M. (2003). Fungal bioreactors: application to white-rot fungi. Reviews in Environmental Science and Biotechnology, 2, 247–259.CrossRefGoogle Scholar
  6. 6.
    Claus, H. (2004). Laccases: structure, reactions, distribution. Micron, 35, 93–96.CrossRefGoogle Scholar
  7. 7.
    Mayer, A. M., & Staples, R. C. (2002). Laccase: new function for an old enzyme. Phytochemistry, 60, 551–565.CrossRefGoogle Scholar
  8. 8.
    Riva, S. (2006). Laccases: blue enzymes for green chemistry. Trends in Biotechnology, 24, 219–226.CrossRefGoogle Scholar
  9. 9.
    Rodriguez Couto, S., & Toca Herrera, J. L. (2006). Industrial and biotechnological application of laccases: a review. Biotechnology Advances, 24, 500–513.CrossRefGoogle Scholar
  10. 10.
    Bollag, J.-M., & Leonowicz, A. (1984). Comparative studies of extracellular fungal laccases. Applied Microbiology and Biotechnology, 48, 849–854.Google Scholar
  11. 11.
    Revankar, S. M., & Lele, S. S. (2006). Increased production of extracellular laccase by the white rot fungus Coriolus versicolor. World Journal of Microbiology and Biotechnology, 22, 921–926.CrossRefGoogle Scholar
  12. 12.
    Jang, M. Y., Ryu, W. R., & Cho, M. H. (2002). Laccase production from repeated batch culture using free mycelia of Trametes sp. Enzyme and Microbial Technology, 30, 741–746.CrossRefGoogle Scholar
  13. 13.
    Arora, D. S., & Gill, P. K. (2000). Laccase production by some white rot fungi under different nutritional conditions. Bioresource Technology, 72, 283–285.CrossRefGoogle Scholar
  14. 14.
    Rosales, E., Rodriguez Couto, S., & Sanroman, A. (2002). New uses of food waste: application to laccase production by Trametes hirsuita. Biotechnology Letters, 24, 701–704.CrossRefGoogle Scholar
  15. 15.
    Kahraman, S. S., & Gurdal, I. H. (2002). Effect of synthetic and natural culture media on laccase production by white rot fungi. Bioresource Technology, 82, 215–217.CrossRefGoogle Scholar
  16. 16.
    Songulashvili, G., Elisashvili, V., Wasser, S. P., Nevo, E., & Hadar, Y. (2007). Basidiomycetes laccase and manganese peroxidase activity in submerged fermentation of food industry wastes. Enzyme and Microbial Technology, 41, 57–61.CrossRefGoogle Scholar
  17. 17.
    Xavier, A. M. R. B., Tavares, A. P. M., Ferreira, R., & Amado, F. (2007). Trametes versicolor growth and laccse induction with by-products of pulp and paper industry. Electronic Journal of Biotechnology, 10, 444–451.CrossRefGoogle Scholar
  18. 18.
    Weuster-Botz, D., Pramatorova, V., Spassov, G., & Wandrey, C. (1995). Use of a genetic algorithm in the development of a synthetic growth medium for Arthrobacter simplex with high hydrocortisone Δ1 – dehydrogenase activity. Journal of Chemical Technology and Biotechnology, 64, 386–392.CrossRefGoogle Scholar
  19. 19.
    Weuster-Botz, D., & Wandrey, C. (1995). Medium optimization by genetic algorithm for continuous production of formate dehydrogenase. Process Biochemistry, 30, 563–571.Google Scholar
  20. 20.
    Findrik, Z., Zelić, B., Bogdan, S., & Vasić-Rački, Đ. (2004). Model-based and experimental optimization using genetic algorithm. Chemical and Biochemical Engineering Quarterly, 18, 105–116.Google Scholar
  21. 21.
    Tišma, M., Žnidaršič-Plazl, P., Plazl, I., Vasić-Rački, Đ., & Zelić, B. (2010). Oxidation of coniferyl alcohol catalyzed by laccases from Trametes versicolor. Acta Chimica Slovenica, 57, 110–117.Google Scholar
  22. 22.
    Tišma, M., Sudar, M., Vasić-Rački, Đ., & Zelić, B. (2010). Mathematical model for Trametes versicolor growth in submerged cultivation. Bioprocess and Biosystems Engineering, 33, 749–758.CrossRefGoogle Scholar
  23. 23.
    Henriquez, C., & Lissi, E. (2002). Evaluation of the extinction coefficient of the ABTS derived radical cation. Boletín de la Sociedad Chilena de Química, 47, 563–566.CrossRefGoogle Scholar
  24. 24.
    Rodríguez Couto, S., Gundín, M., Lorenzo, M., & Sanromán, A. (2002). Screening of supports for laccase production Trametes versicolor in semi-solid-state conditions. Determination of optimal operation conditions. Process Biochemistry, 38, 249–255.CrossRefGoogle Scholar
  25. 25.
    Rancano, G., Lorenzo, M., Morales, N., Rodriquez Couto, S., & Sanroman, M. A. (2003). Production of laccase by Trametes versicolor in an airlift fermentor. Process Biochemistry, 39, 467–473.CrossRefGoogle Scholar
  26. 26.
    Pazarlioğlu, N. K., Sariişik, M., & Telefoncu, A. (2005). Laccase: Production by Trametes versicolor and application to denim washing. Process Biochemistry, 40, 1673–1678.CrossRefGoogle Scholar
  27. 27.
    Aktas, N., Cicek, H., Unal, A. T., Kibarer, G., Kolankaya, N., & Tanyolac, A. (2001). Reaction kinetics for laccase-catalyzed polymerization of 1-naphthol. Bioresource Technology, 80, 29–36.CrossRefGoogle Scholar
  28. 28.
    Tavares, A. P. M., Coelho, M. A. Z., Coutinho, J. A. P., & Xavier, A. M. R. B. (2005). Laccase improvement in submerged cultivation: induced production and kinetic modelling. Journal of Chemical Technology and Biotechnology, 80, 669–676.CrossRefGoogle Scholar
  29. 29.
    Tavares, A. P. M., Coelho, M. A. Z., Agapito, M. S. M., Coutinho, J. A. P., & Xavier, A. M. R. B. (2006). Optimization and modeling of laccase production by Trametes versicolor in a bioreactor using statistical experimental design. Applied Microbiology and Biotechnology, 134, 233–248.Google Scholar
  30. 30.
    Lorenzo, M., Moldes, D., Rodríguez Couto, S., & Sanromán, A. (2002). Improving laccase production by employing different lignocellulosic wastes in submerged cultures of Trametes versicolor. Bioresource Technology, 82, 109–113.CrossRefGoogle Scholar
  31. 31.
    Elisashvili, V., & Kachlishvili, E. (2008). Effect of growth substrate, method of fermentation, and nitrogen source on lignocellulose-degrading enzymes production by white-rot basidiomycetes. Journal of Industrial Microbiology and Biotechnology, 35, 1531–1538.CrossRefGoogle Scholar
  32. 32.
    Saparrat, M. C. N., Arambarri, A. M., & Balatti, P. (2007). Growth and extracellular laccase production in liquid cultures of Minimidochium parvum LPSC # 548 strain. Bulletin of the Botanical Society of Argentina, 42, 39–47.Google Scholar
  33. 33.
    Malarczyk, E., Jarosz-Wilkolazka, A., & Kochmanska-Rdest, J. (2003). Effect of low doses of guaiacol and ethanol on enzymatic activity of fungal cultures. Nonlinearity of Biology, Toxicology, and Medicine, 1, 167–168.CrossRefGoogle Scholar
  34. 34.
    Žnidaršič-Plazl, P., & Plazl, I. (2010). Development of a continuous steroid biotransformation process and product extraction within microchannel system. Catalysis Today, 157, 315–320.CrossRefGoogle Scholar
  35. 35.
    Kaup, B.-A., Ehrich, K., Pescheck, M., & Schrader, J. (2008). Microparticle-enhanced cultivation of filamentous microorganisms: increased chloroperoxidase formation by Caldariomyces fumago as an example. Biotechnology and Bioengineering, 99, 491–498.CrossRefGoogle Scholar
  36. 36.
    Driouch, H., Sommer, B., & Wittmann, C. (2010). Morphology engineering of Aspergillus niger for improved enzyme production. Biotechnology and Bioengineering, 105, 1059–1068.Google Scholar
  37. 37.
    Žnidaršič-Plazl, P. (2006). The influence of some engineering variables upon the morphology of Rhizopus nigricans in a stirred tank bioreactor. Chemical and Biochemical Engineering Quarterly, 20, 275–280.Google Scholar

Copyright information

© Springer Science+Business Media, LLC 2011

Authors and Affiliations

  • Marina Tišma
    • 1
  • Polona Žnidaršič-Plazl
    • 2
  • Đurđa Vasić-Rački
    • 3
  • Bruno Zelić
    • 3
    Email author
  1. 1.Faculty of Food Technology OsijekJ. J. Strossmayer University of OsijekOsijekCroatia
  2. 2.Faculty of Chemistry and Chemical TechnologyUniversity of LjubljanaLjubljanaSlovenia
  3. 3.Faculty of Chemical Engineering and TechnologyUniversity of ZagrebZagrebCroatia

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