Improvement of Fungal Cellulase Production by Solid State Fermentation

Author(s): Hind Leghlimi, Ilhem Djezzar-Mihoubi, Hayet Boukhalfa-Lezzar, Scheherazad Dakhmouche, Leila Bennamoun, Zahia Meraihi

Cellulase production studies have been carried out using the fungal strains Trichoderma longibrachiatum and Aspergillus terreus by using two different lignocellulosic materials for solid state fermentation (SSF). The effect of basic fermentation parameters (substrate, moisture content and the fermentation time) on enzyme production was studied. Maximum cellulase production (FPA and endoglucanase) of Trichoderma longibrachiatum were 33.83 U/gds and 167.4 U/gds, respectively, which represented a 1.75 fold improved activities than that of Aspergillus terreus (21.5 U/gds and 95.82 U/gds, respectively) using wheat bran as substrate. The optimal conditions for cellulase production with wheat bran were found to be: initial moisture content 70% and the optimal incubation time for production was three days. The endoglucanase of Trichoderma longibrachiatum was thermostable and showed a half-life of 1 hour at 70°C, while the filter paper activity (APF) lost its total activity after 1 hour at 70°C. Results indicate the scope for further optimization of the production conditions to obtain higher cellulase titres using the Trichoderma longibrachiatum strain under SSF.

Cellulase, Trichoderma longibrachiatum, Aspergillus terreus, Solid state fermentation, Lignocellulosic material, Wheat bran

1. Abdullah, J. J., Greetham, D., Pensupa, N., Tucker, G. A., Du, C. (2016). Optimizing Cellulase Production from Municipal Solid Waste (MSW) using Solid State Fermentation (SSF). J Fundam Renewable Energy Appl, 6, 206. http://dx.doi.org/10.4172/2090-4541.1000206.
2. Bhat, M. K., Bhat, S. (1997). Cellulose degrading enzymes and their potential industrial applications. Biotechnol Adv, 15(3-4):583-620.
3. Busto, M. D., Ortega, N., Perez-Mateos, M. (1996). Location, Kinetics and stability of cellulases induced in Trichoderma reesei cultures. Bioresour Technol, 57, 187-192.
4. Dedavid, E. S. L. A., Lopes, F. C., Silveira, S. T., Brandelli, A. (2008). Production of cellulolytic enzymes by Aspergillus phoenicis in grape waste using response surface methodology. Appl Biochem Biotechnol, http://dx.doi.org/10.1007/s12010-008-8190-7.
5. Devendra, P., Maurya, D. S., Durgesh, P., Maurya, J. P. (2012). Optimization of solid state fermentation conditions for the production of cellulase by Trichoderma reesei. J Environ Biol, 33, 5-8.
6. Duff, S. J., Murray, W.D. (1996). Bioconversion of forest products industry waste cellulosics to fuel ethanol. Bioresour Technol, 55, 1-33.
7. Durand, H., Clanet, M., Tiraby, G. (1988). Genetic improvement of Trichoderma reesei for large scale cellulase production. Enzyme Microb Technol, 10, 341-6.
8. FAOSTAT. (2012). FAOSTAT Agriculture Data, Food and Agriculture Organization of the United Nations, http://faostat.fao.org/site/339/default.aspx (accessed August).
9. Gao, J., Weng, H., Zhu, D., Yuan, M., Guan, F., Xi, Y. (2008). Production and characterization of cellulolytic enzymes from the thermoacidophilic fungal Aspergillus terreus M11 under solid-state cultivation of corn stover. Bioresour Technol, 99, 7623–7629.
10. Ghose, T. K. (1987). Measurement of cellulase activities. Pure Appl Chem, 59,257-268.
11. Gong, W., Zhang, H., Liu, S., Zhang, L., Gao, P., Chen, G., Wang, L. (2015). Comparative secretome analysis of Aspergillus niger, Trichoderma reesei, and Penicillium oxalicum during solid-state fermentation. Appl Biochem Biotechnol. http://dx.doi.org/10.1007/s12010-015-1811-z.
12. Hagerdal, B., Ferchak, J. D., Pye, E. K. (1980). Saccharification of cellulose by the cellulolytic enzyme system of Thermomonospora sp.I. Stability of cellulolytic activities with respect to time, temperature, and pH. Biotechnol Bioeng, 22, 1515-1526.
13. Horn, S. J., Vaaje-Kolstad, G., Westereng, B., Eijsink, V. G. (2012). Novel enzymes for the degradation of cellulose. Biotechnology for Biofuels, 5, 45. http://dx.doi.org/10.1186/1754-6834- 5-45
14. Ikram-ul-Haq, M. M. J., Khan, T. S. (2006). An innovative approach for hyper production of cellulolytic and hemicellulolytic enzymes by consortium of Aspergillus niger MSK-7 and Trichoderma viride MSK-10. Afr J Biotechnol, 5, 609-614.
15. Kamm, B., Kamm, M. (2004). Principles of biorefineries. Appl Microbiol Biotechnol, 64(2), 137-145.
16. Kang, S.W., Park, Y. S., Lee, J. S., Hong, S. I., Kim, S. W. (2004). Produciton of cellulases and hemicellulases by Aspergillus niger KK2 from lignocellulosic biomass. Bioresour Technol, 91, 153-156.
17. Kaur, B., Oberoi, H., Chadha, B. S. (2014). Enhanced cellulase producing mutants developed from heterokaryotic Aspergillus strain. Bioresource Technology, 156, 100-107. http://dx.doi.org/10.1016/j.biortech.2014.01.016
18. Lynd, L. R., Weimer, P. J., Van Zyl, W. H., Pretorius, I. S. (2002). Microbial cellulose utilization: fundamental and biotechnology. Microbiology and Molecular Biology, 66(3), 506-507. http://dx.doi.org/10.1128/MMBR.66.3.506-577.
19. Maes, C., Delcour, J. A. (2001). Alkaline hydrogen peroxide extraction of wheat bran non-starch polysaccharides. J Cereal Sci, 34(1), 29-35.
20. Margaritis, A., Merchant, R. F. (1986). Optimization of fermentation conditions for thermostable cellulase production by Thielavia terrestris. J Ind Microbiol, 1, 149-156.
21. Maurya, D. P., Singh, D., Pratap, D., Maurya, J. P. (2012). Optimization of solid state fermentation conditions for the production of cellulase by Trichoderma reesei. J Environ Biol, 33, 5-8.
22. Mekala, N. K., Singhania, R. R., Sukumaran, R. K., Pandey, A. (2008). Cellulase production under solid-state fermentation by Trichoderma reesei RUT C30: statistical optimization of process parameters. Appl Biochem Biotechnol, 151, 122-131.
23. Miller, G.L. (1959). Use of dinitrosalicylic acid reagent for determination of reducing sugar. Anal Chem, 31, 426-428.
24. Mrudula, S., Murugammal, R. (2011). Production of cellulase by Aspergillus niger under submerged and solid state fermentation using coir waste as a substrate. Braz J Microbiol, 42, 1119-1127.
25. Murray, P., Aro, N., Collins, C., Grassick, A., Penttila, M., Saloheimo, M., Tuohy, M. (2004). Expression in Trichoderma reesei and characterization of a thermostable family 3β-glucosidase from the moderately thermophilic fungus Talaromyces emersonii. Protein Expr Purif, 38(2), 248-257.
26. Narra, M., Dixit, G., Divecha, J., Kumar, K., Madamwar, D., Shah, A. R. (2014). Production, purification and characterization of a novel GH12 family endoglucanase from Aspergillus terreus and its application in enzymatic degradation of delignified rice straw. International journal of Biodeterioration and Biodegradation, 88, 150-161. http://dx.doi.org/10.1016/j.ibiod.2013.12.016
27. Narra, M., Dixit, G., Divecha, J., Madamwar, D., Shah, A. R. (2012). Production of Cellulases by Solid State Fermentation with Aspergillus terreus and Enzymatic Hydrolysis of Mild Alkali-Treated Rice Straw. Bioresource Technology, 121, 355-361. http://dx.doi.org/10.1016/j.biortech.2012.05.140
28. Ogel, Z. B., Yarangumeli, K., Du, H., Ifrij, J. (2001). Submerged cultivation of Scytalidium thermophilum on complex lignocellulosic biomass. Enzyme Micobiol Technol, 28, 689-695.
29. Pandey, A. (2003). Solid-state fermentation. Biochem Eng J, 13(2-3), 81-84.
30. Singhania, R. R., Sukumaran, R. K., Patel, A. K., Larroche, C., Pandey, A. (2010). Advancement and comparative profiles in the production technologies using solid state and submerged fermentation for microbial cellulases. Enzyme Microb Technol, 46 (7), 541-549.
31. Singhania, R. R., Sukumaran, R. K., Pillai, A., Prema, P., Szakas, G., Pandey, A. (2006). Solide state fermentation of lignocellulosic substrates for cellulase production by Trichoderma reesei NRRL 11460. Indian Journal of Biotechnology, 5, 332-336.
32. Sohail, M., Siddiqi, R., Ahmad, A., Khan, S. A. (2009). Cellulase production from Aspergillus niger MS82: effect of temperature and pH. New Biotechnol, 25, 437-41.
33. Soni, R., Nazir, A., Chadha, B. S., Saini, H. S. (2008). Novel sources of fungal cellulases for efficient deinking of composite paper waste. Bioresources, 3, 234-246.
34. Sun, H., Ge, X., Hao, Z., Peng, M. (2010). Cellulase production by Trichoderma sp. on apple pomace under solid state fermentation. African Journal of Biotechnology, 9 (2), 163-166.
35. Vu, V. H., Pham, T.A., Kim, K. (2010). Improvement of a fungal strain by repeated and sequential mutagenesis and optimization of solid-state fermentation for the hyper-production of rawstarch-digesting enzyme. J Microbiol Biotechnol, 20, 718-26.
36. Vu, V. H., Pham, T. A., Kim K. (2011). Improvement of Fungal Cellulase Production by Mutation and Optimization of Solid State Fermentation. Mycobiology, 39(1), 20-25.