Production of Glucoamylase from Aspergillus Awamori Nrrl-356 using 2-Factorial Plackett-Burman Design
[1]
Sikander Ali , Institute of Industrial Biotechnology (IIB), GC University Lahore, Pakistan.
[2]
Mian Sahib Zar , Institute of Industrial Biotechnology (IIB), GC University Lahore, Pakistan; Centre of Excellence in Molecular Biology, University of the Punjab Lahore, Pakistan.
[3]
Nazia Zafar , Institute of Industrial Biotechnology (IIB), GC University Lahore, Pakistan.
[4]
Ahmad Ali Shahid , Centre of Excellence in Molecular Biology, University of the Punjab Lahore, Pakistan.
In the present study, we report on the production of glucoamylase by Aspergillus awamori NRRL-356 under solid-state fermentation (SSF). A range of different agricultural by-products such as corn cobs, bagasse, soybean meal, wheat straw, wheat bran and mustard cake were evaluated. Ungrinded wheat straw at a level of 10g in 250 ml Erlenmeyer flasks was evaluated as the superlative substrate. Cultural conditions were optimized in order to further enhance the enzyme yield. The maximum enzyme activity (63.54±3.55 U/g with 450±16 µg/ml total protein content) was achieved when substrate was moistened with distilled water at the level of 12.5 ml i.e., at 1.25:1 ratio. More notably, again distilled water at the level of 150 ml was found as the best extracting agent. The significant improvement in enzyme production was attributed to the process parameters including incubation temperature (30ºC), time of fermentation (72 h) and size of inoculum (10%) when the selected fungal culture was further identified using the 2-factorial Plackett-Burman Design (PBD). Further more, the application of response surface methodology (RSM) was proved to be crucial in achieving a high performance batch fermentation process for glucoamylase production, which is highly significant (HS, p≤0.05, LSD~0.216). This is a kind of novel study which has not been reported earlier, however, scale up studies is pre-requisite for the commercial exploitation of fungal culture.
Aspergillus awamori, Glucoamylase, Solid-State Fermentation, 2-Factorial Plackett-Burman Design, Agricultural By-Products
[1]
Lee J and Paetzel M, Structural Biology and Crystallization Communications, Acta Cryst, 67 (2011) 188-192.
[2]
Zambare V, Solid State Fermentation of Aspergillus oryzae for Glucoamylase Production on Agro residues, Int. J. Life Sci, 4 (2010) 16-25.
[3]
Zaroog M S and Saad T, Formation of molten globule-like state during acid denaturation of Aspergillus niger glucoamylase, Proc. Biochem, 47 (2012) 775–784.
[4]
Sun H, Zhao P, Ge X, Xia Y, Hao Z, Liu J and Peng M, Recent advances in microbial raw starch degrading enzymes, Appl. Biochem. Biotechnol, 160 (2010) 988–1003.
[5]
Bertolin T E, Schmidell W, Maiorano A E, Casara J and Costa J A, Influence of carbon, nitrogen and phosphorous sources on glucoamylase production by Aspergillus spp. in solid state fermentation. Z. Naturforsch, 58 (2003) 708–712.
[6]
Negi S and Rintu B, Optimization of culture parameters to enhance production of amylase and protease from Aspergillus awamori in a single fermentation. Afr. J. Biochem Res, 4(3) (2010) 73-80.
[7]
Norouzian D, Akbarzadeh A, Scharer J M and Young M, Fungal glucoamylase. Biotechnol Adv, 24 (2006) 80–85.
[8]
Dojnov B and Zoran V, Fast and reliable method for simultaneous zymographic detection of glucoamylase and a-amylase in fungal fermentation. Analytic. Biochem, 421 (2012) 802–804.
[9]
Kareem S O, Akpan I and Oduntan S B, Cowpea waste: A novel substrate for solid state production of amylase by Aspergillus oryzae, Afr. J. Microbiol. Res, 3(12) (2009) 974-977.
[10]
Vu V H, Tuan A P and Keun K, Improvement of a Fungal Strain by Repeated and Sequential Mutagenesis and Optimization of Solid-State Fermentation for the Hyper-Production of Raw-Starch-Digesting Enzyme. J. Microbiol. Biotechnol, 20(4) (2010) 718–726.
[11]
Arnthong J, Boonpa W, Kenji S, Jean-Jacques S and Vichien K, Statistical screening of factors affecting glucoamylase production by thermotolerant Rhizopus microspores TISTR 3518 using Plackett-Burman design. Afr. J. Biotechnol, 9(43) (2010) 7312-7316.
[12]
Akoh C C, Chang S W, Lee G C and Shaw J F, Biocatalysis for the production of industrial products and functional foods from rice and other agricultural products, J. Agric. Food Chem, 56 (2008) 10445–10451.
[13]
Burkert J F M, Kalil S J, Filho F M and Rodrigues M I, Parameters optimization for enzymatic assays using experimental design, Braz. J. Chem. Engin, 23 (2006) 163–170.
[14]
Gracheva I M, Gernet M V, Gavrilova N N and Razarenova N F, Effect of some amino acids on the biosynthesis of glucoamylase by cultured Endomycopsis species, Appl. Biochem. Microbiol, 14 (1978) 871–877.
[15]
Ahuja, S K, Ferreira G M and Morreira, A R, Application of Plackett-Burman design and response surface methodology to achieve exponential growth of aggregated shipworm bacterium. Biotechnol. Bioeng, 85 (2004) 666–675.
[16]
Kumar P and Satyanarayana T, Microbial glucoamylases: characteristics and application, Crit. Rev. Biotechnol, 29 (2009) 225–255.
[17]
Caldwell K D, Roff A, Margereta B and Jerker P, Estimation of glucoamylase. Biotechnol. Bioengin, 18 (1968) 1592.
[18]
Nelsons N, A photometric adaptation of the Somogyi method for the determination of glucose, J. Biol. Chem, 153 (1944) 375–380.
[19]
Smogyi M, Notes on sugar determination, J. Biol. Chem, 195 (1937) 19–23.
[20]
Bradford N M, A rapid and sensitive method for quantization of microorganisms’ qualities of protein utilizing the principle of protein dye binding analysis. Biochem, 72 (1976) 248–254.
[21]
Snedecor G W and Cochran W G, Statistical Methods. 7th Edition, Ames, lowa State Univ. Press, lowa, USA, (1980).
[22]
Peixoto-Nogueira S C, Sandrim V C, Guimaraes L H, Jorge J A, Terenzi, H F and Polizeli M L, Evidence of thermostable amylolytic activity from Rhizopus microsporus var. Rhizopodiformis using wheat bran and corncob as alternative carbon source, Bioproc. Biosyst. Eng, 31 (2008) 329–334.
[23]
Aghdam J J, Khajeh K, Ranjbar B and Gorgani M N, Production of glucoamylase from Aspergillus niger, effects on structure, activity and stability, Braz. J. Microbiol, 39 (2005) 1517–1532.
[24]
Makhamud S A, Mashkhur V A, Khaddad M E and Gammal S E, Effect of the composition of the medium and the conditions of Aspergillus foetidus cultivation on the biosynthesis of glucoamylase, Microbiol, 47 (1978) 220–225.
[25]
Mala J G, Edwinoliver N G, Kamini N R and Puvanakrishnan R, Mixed substrate solid state fermentation for production and extraction of glucoamylase from Aspergillus niger MTCC 2594, Gen. Appl. Microbiol. 53 (2007) 247–253.
[26]
Silveira S, Oliveira T, Costa M S and Kalil S J, Optimization of glucoamylase production by Aspergillus niger in solid-state fermentation, Appl. Biochem. Biotechnol, 128 (2006) 131–140.
[27]
Ghosh A, Chatterjee B, Das A, Induction and catabolite repression of high-affinity glucoamylase in Aspergillus terreus strain 4. Engin. Microbiol, 136 (1990) 1307–1311.
[28]
Anto H, Trivedi U B, Patel K C, Glucoamylase production by solid-state fermentation using rice flake manufacturing waste products as substrate, Bioresourc. Technol, 97 (2006) 1161–1166.
[29]
Stamford T L M, Stamford N P, Coelho L C B B and Araujo J M, Production and characterization of a thermostable glucoamylase from Streptosporangium endophyte of maize leaves, Bioresourc. Technol, 83 (2002) 105–109.
[30]
Satyanarayana T, Noorwez S M, Kumar S, Rao J L, Ezhilvannan M and Kaur P, Development of an ideal starch saccharification process using amylolytic enzymes from thermophiles, Biochem. Soc. Trans, 32 (2004) 276–278.
[31]
Jin B, Leeuwen H J V, Patel B and Young Q Y M, Utilization of starch processing wastewater for production of glucoamylase and fungal α-amylase by Aspergillus oryzae, Environ. Engin, 6 (1998) 179–189.
[32]
Mamo G and Gessesse A, Production of raw-starch digesting glucoamylase by Aspergillus sp. GP-21 in solid state fermentation, J. Microbiol. Biotechnol, 22 (1999) 622–626.
