Cellulase-producing bacteria were isolated from soil and identified as Pseudomonas fluorescens, Bacillus subtilIs, E. coli, and Serratia marcescens. Optimization of the fermentation medium for maximum cellulase production was carried out. The culture conditions like pH, temperature, carbon sources, and nitrogen sources were optimized. The optimum conditions found for cellulase production were 40°C at pH 10 with glucose as carbon source and ammonium sulphate as nitrogen source, and coconut cake stimulates the production of cellulase. Among bacteria, Pseudomonas fluorescens is the best cellulase producer among the four followed by Bacillus subtilis, E. coli, and Serratia marscens. 1. Introduction Cellulose is the most abundant biomass on Earth [1]. It is the primary product of photosynthesis in terrestrial environments and the most abundant renewable bioresource produced in the biosphere [2, 3]. Cellulose is commonly degraded by an enzyme called cellulase. This enzyme is produced by several microorganisms, commonly by bacteria and fungi [4–7]. Cellulose is the principal constituent of the cell wall of most terrestrial plants. The source of cellulose is in plants and it is found as microfibrils (“2–20？nm” in diameter and “100–40,000？nm” long). These form the structurally strong framework in the cell walls. Despite a worldwide and enormous utilization of natural cellulosic sources, there are still abundant quantities of cellulosic sources and there are still abundant quantities of cellulose containing raw materials and waste products that are not exploited or which could be used more efficiently. The problem in this respect is, however, to develop processes that are economically profitable. Complete hydrolysis of the enzyme requires synergistic action of 3 types of enzymes, namely, cellobiohydrolase, endoglucanase or carboxymethylcellulase (CMCase), and beta-glucosidases [8]. Bacteria which have high growth rate as compared to fungi have good potential to be used in cellulase production. However, the application of bacteria in producing cellulase is not widely used. The cellulolytic property of some bacterial genera such as Cellulomonas, Cellvibrio, Pseudomonas sp [9]. Bacillus, and Micrococcus [7], was also reported. Enzyme production is closely controlled in microorganisms and for improving its productivity these controls can be ameliorated. Cellulase yields appear to depend upon a complex relationship involving a variety of factors like inoculums size, pH value, temperature, presence of inducers, medium additives, aeration, growth time, and so forth [7].
References
[1]
P. Tomme, R. A. J. Warren, and N. R. Gilkes, “Cellulose hydrolysis by bacteria and fungi,” Advances in Microbial Physiology, vol. 37, pp. 1–81, 1995.
[2]
M. Jarvis, “Cellulose stacks up,” Nature, vol. 426, no. 6967, pp. 611–612, 2003.
[3]
Y.-H. P. Zhang and L. R. Lynd, “Toward an aggregated understanding of enzymatic hydrolysis of cellulose: Noncomplexed cellulase systems,” Biotechnology and Bioengineering, vol. 88, no. 7, pp. 797–824, 2004.
[4]
A. H. Bahkali, “Influence of various carbohydrates on xylanase production in Verticillium tricorpus,” Bioresource Technology, vol. 57, no. 3, pp. 265–268, 1996.
[5]
P. Magnelli and F. Forchiassin, “Regulation of the cellulase complex production by Saccobolus saccoboloides: induction and repression by carbohydrates,” Mycologia, vol. 91, no. 2, pp. 359–364, 1999.
[6]
C. S. Shin, J. P. Lee, J. S. Lee, and S. C. Park, “Enzyme production of Trichoderma reesei rut C-30 on various lignocellulosic substrates,” Applied Biochemistry and Biotechnology A, vol. 84–86, pp. 237–245, 2000.
[7]
G. Immanuel, R. Dhanusha, P. Prema, and A. Palavesam, “Effect of different growth parameters on endoglucanase enzyme activity by bacteria isolated from coir retting effluents of estuarine environment,” International Journal of Environmental Science and Technology, vol. 3, no. 1, pp. 25–34, 2006.
[8]
M. K. Bhat, “Cellulases and related enzymes in biotechnology,” Biotechnology Advances, vol. 18, no. 5, pp. 355–383, 2000.
[9]
K. Nakamura and K. Kppamura, “Isolation and identification of crystalline cellulose hydrolysing bacterium and its enzymatic properties,” Journal of Fermentation Technology, vol. 60, no. 4, pp. 343–348, 1982.
[10]
Y. J. Lee, B. K. Kim, B. H. Lee et al., “Purification and characterization of cellulase produced by Bacillus amyoliquefaciens DL-3 utilizing rice hull,” Bioresource Technology, vol. 99, no. 2, pp. 378–386, 2008.
[11]
S. Saha, R. N. Roy, S. K. Sen, and A. K. Ray, “Characterization of cellulase-producing bacteria from the digestive tract of tilapia, Oreochromis mossambica (Peters) and grass carp, Ctenopharyngodon idella (Valenciennes),” Aquaculture Research, vol. 37, no. 4, pp. 380–388, 2006.
[12]
Y. Nishida, K. I. Suzuki, Y. Kumagai, H. Tanaka, A. Inoue, and T. Ojima, “Isolation and primary structure of a cellulase from the Japanese sea urchin Strongylocentrotus nudus,” Biochimie, vol. 89, no. 8, pp. 1002–1011, 2007.
[13]
J. Pranner, “Environmental Microbiology and Waste Utilization,” in G/AMV Proceedings, Emejuaiwe, Ed., pp. 67–69, Academic Press, London, UK, 1979.
[14]
M. Camassola and A. J. P. Dillon, “Production of cellulases and hemicellulases by Penicillium echinulatum grown on pretreated sugar cane bagasse and wheat bran in solid-state fermentation,” Journal of Applied Microbiology, vol. 103, no. 6, pp. 2196–2204, 2007.
[15]
C. Koomnok, “Selection of cellulase producing thermophilic fungi,” in Proceedings of the 31st Congress on Science and Technology of Thailand of Technology, Suranaree University, October 2005.
[16]
J. R. Cherry and A. L. Fidants, “Directed evolution of industrial enzymes: an update,” Current Opinion in Biotechnology, vol. 14, no. 4, pp. 438–443, 2003.
[17]
K. Apun, B. C. Jong, and M. A. Salleh, “Screening and isolation of a cellulolytic and amylolytic Bacillus from sago pith waste,” Journal of General and Applied Microbiology, vol. 46, no. 5, pp. 263–267, 2000.
[18]
G. L. Miller, “Use of dinitrosalicylic acid reagent for determination of reducing sugar,” Analytical Chemistry, vol. 31, no. 3, pp. 426–428, 1959.
[19]
R. E. Buchanan and N. E. Gibbons, Bergey's of Determinative Bacteriology, Williams & Wilkins Co., Philadelphia, PA, USA, 1974.
[20]
P. Chantawannakul, A. Oncharoen, K. Klanbut, E. Chukeatirote, and S. Lumyong, “Characterization of cellulases of Bacillus subtilis strain 38 isolated from traditionally fermented soybean in northern Thiland,” ScienceAsia, vol. 28, pp. 241–245, 2002.
[21]
A. M. Abdel-Mawgoud, M. M. Aboulwafa, and N. A. H. Hassouna, “Optimization of surfactin production by bacillus subtilis isolate BS5,” Applied Biochemistry and Biotechnology, vol. 150, no. 3, pp. 305–325, 2008.
[22]
W. Win, Z. Lianhui, L. Dog, W. Yong, Z. Zhenshan, and M. Zhihuai, “Conditions study of cellulose and acid protease production during the process of solid state fermentation of flaxseed meal,” American Society of Agriculture and Biological Engin, vol. 34, no. 6, pp. 45–51, 2008.
[23]
E. Jansová, Z. Schwarzová, and J. Chaloupka, “Sporulation and synthesis of extracellular proteinases in Bacillus subtilis are more temperature-sensitive than growth,” Folia Microbiologica, vol. 38, no. 1, pp. 22–24, 1993.
[24]
M. K. Bakare, I. O. Adewale, A. Ajayi, and O. O. Shonukan, “Purification and characterization of cellulase from the wild-type and two improved mutants of Pseudomonas fluorescens,” African Journal of Biotechnology, vol. 4, no. 9, pp. 898–904, 2005.
[25]
A. K. Ray, A. Bairagi, K. Sarkar Ghosh, and S. K. Sen, “Optimization of fermentation conditions for cellulase production by Bacillus subtilis CY5 and Bacillus circulans TP3 isolated from fish gut,” Acta Ichthyologica et Piscatoria, vol. 37, no. 1, pp. 47–53, 2007.
[26]
M. Ishihara, M. Matsunaga, N. Hayashi, and V. Ti？ler, “Utilization of D-xylose as carbon source for production of bacterial cellulose,” Enzyme and Microbial Technology, vol. 31, no. 7, pp. 986–991, 2002.
[27]
K. Toda, T. Asakura, M. Fukaya, E. Entani, and Y. Kawamura, “Production of cellulose from D-arabitol by Acetobacter xylinum KU-1,” Bioscience, Biotechnology, and Biochemistry, vol. 59, no. 8, pp. 1564–1565, 1995.
[28]
K. V. Ramana, A. Tomar, and L. Singh, “Effect of various carbon and nitrogen sources on cellulose synthesis by Acetobacter xylinum,” World Journal of Microbiology and Biotechnology, vol. 16, no. 3, pp. 245–248, 2000.
[29]
M. Mandels, “Microbial source of cellulose,” Biotechnology and Bioengineering, vol. 5, pp. 81–105, 1975.
