International Journal of Biology Research  23    International Journal of Biology Research ISSN: 2455-6548, Impact Factor: RJIF 5.22 www.biotechjournals.com Volume 1; Issue 2; May 2016; Page No. 23-27 Production of Cellulase by Aspergillus niger NCFT 4263.10 using Agro waste as a Substrate 1 Bijay Kumar Sethi, 2 Asutosh Shyamaprasad Dash, 3 Amrita Satapathy, 4 Subodh Kumar Tripathy, 5 Sidharth Parida, 6 Bikash Chandra Behera 1, 2, 3, 4, 5 Department of Biotechnology, MITS School of Biotechnology, Bhubaneswar-751024, Odisha, India 2 Department of Biotechnology, North Orissa University, Baripada-757003, Odisha, India Abstract Aspergillus niger NCFT 4263.10 was evaluated for the production of cellulase by liquid static surface fermentation (LSSF), Liquid shaking fermentation (LShF) and solid state fermentation (SSF). Higher cellulase and protein secretion was observed when fermentation carried out in LSSF condition as compared to ShF and SSF. Maximum cellulase production was observed in LSSF (393.5±3.41U/ml) when wheat bran was used as the substrate followed by banana peels and saw dust. Immobilized spores of A. niger applied for fermentation up to five cycle and study revealed that when wheat bran was used as the carbon source, the activity of cellulase was found to be maximum. Highest Cellulase activity observed at second cycle (415.4±6.5U/ml) followed by third cycle (379.24 ± 2.65U/ml), fourth cycle (340.07± 2.6U/ml) and fifth cycle (204.25± 1.71U/ml). Keywords: Aspergillus niger, banana peels, fermentation, wheat bran 1. Introduction Many agricultural by products from agricultural activities and agro based processing litter the environment and constitute waste problems [1] . Large amounts of these agricultural residues are also frequently burnt in the fields causing severe environmental pollution by increase in the CO2 level [2] . Efficient utilization and microbial biodegradation of, large quantities of agro-industrial wastes and crop residues will lead to several processes of great economic value [3] . These agro wastes are mainly composed of lignocellulosic material which gained considerable interest as an abundant renewable bio energy resource because of their possible use in secondary fermentation process for the production of food, fuel and chemicals. For biodegradation and bioconversion of these cellulosic materials into useful product application of microbial cellulase is used as an essential tool for several researchers all over the world. Most of the bacteria and fungi degrade this cellulosic material by producing enzyme called cellulase [4] . Cellulase is the enzyme that hydrolyses the β-1, 4-glycosidic bonds in the polymer to release glucose units and are among the industrially important hydrolytic enzymes of great significance in present day biotechnology [4] . The use of agro wastes as the basis for the cultivation media is a matter of great interest today, aiming to decrease the costs of energy production and meeting the increase in awareness on energy conservation and recycling [1] . Agro wastes such as rice straw, wheat bran, banana peels, corn stover, sugar cane bagasse, pomace, corn cobs etc are used as substrate in solid state fermentation [5] . Cellulase production using these agro wastes by different organisms in submerged fermentation state has received more attention and is found to be cost prohibitive because of high cost of process engineering [6, 7] . More than 14,000 species of fungi have been found to be active in cellulose degradation. Among several hypercellulolytic fungal strains, filamentous fungi such as Trichoderma and Aspergillus sp. are considered as the major microbial sources for production of cellulose-degrading enzymes [8, 9] . Trichoderma sp. is a well-known commercial cellulase producer but lacks sufficient β-glucosidase activity [10] . In contrary, Aspergillus sp. produces high amounts of β- glucosidase in the extracellular medium and is commonly used for commercial production of this enzyme. The present research investigation deals with studies on the production cellulase from Aspergillus niger using different agro waste as a substrate. 2. Materials and Methods 2.1. Sample collection and inoculums preparation Fungal strains, such as, Aspergillus niger (NCFT 4263.10) was used as a microbial strain. Banana peels (BP), saw dust (SD) and wheat bran (WB) were collected from local area. These agro-substrates were dried at 60°C for 48 h or more until the moisture content was reduced. Cooled samples were later grounded in a blender and kept in sterile containers until required [11] . Spore suspension (1 ml) having spore concentration of about 1 × 10 7 cells ml -1 from 7 days old culture was used as inoculum in the experiments. 2.2. Growth medium condition for cellulase production In 150 ml capacity Erlenmeyer flasks, 50 ml of sterilized fermentation medium having individually BP, SD, WB and commercial carboxymethyl cellulose was inoculated with 1.0 × 10 7 cells ml -1 from 7 days old cultures of A. niger and incubated at 30 ± 2°C at liquid static surface fermentation, solid static fermentation and by shaking condition at 100 rpm. Samples were processed at 24 h of regular interval up to 120 h for recovery of cellulase and its activity assay. The fermented broth (50ml) cultures were centrifuged at 10,000 rpm, at 4°C for 10 min to obtain the crude supernatant. Further, the clear crude supernatant was passed through Whatman No. 1 filter paper before cellulase activity assay. The remaining crude enzyme was preserved at -20°C for further use.