Extracellular endo-mannanase from Bacillus sp CFR1601: Economical production using response surface methodology and downstream processing using aqueous two phase system

Bacillus sp. CFR1601, isolated from decaying plant litter, produced an extra-cellular endo-mannanase (198.0 IU/g) under solid state fermentation (SSF) using defatted coconut residue as the prime solid substrate. In order to enhance endo-mannanase production, three component, five level central composite design (CCD) of response surface methodology (RSM) was used. Based on contour plots and variance analysis, optimum conditions for endomannanase production from Bacillus sp. CFR1601 were attained when defatted coconut residue was supplemented with sesame oil meal (10.0, w/w), Tween-80 (0.2%, v/v) and inoculated with bacterial cells from log phase (12 h old; OD600 nm approximate to 3.6). The empirical model developed through RSM brought about 4.04-4.39-fold (800.0-870.0 IU/g) improvement in endo-mannanase yield as compared to un-optimized growth conditions. Downstream processing of endo-mannanase from SSF media was carried out for the first time using polyethylene glycol (PEG)/salt aqueous two phase system (ATPS). ATPS system consisting of a combination of PEG 3350 12.0% (w/w), Na2SO4 12.0% (w/w), protein load 10.0% (w/w) and pH 5.0 resulted in one-sided partitioning of endo-mannanase towards bottom phase with 3.8-fold purification and 95.4% recovery. Second stage ATPS with fresh top phase further improved purification of endo-mannanase to 12.32-fold. Our overall results suggest a cost-effective and integrated process for production and downstream processing of endo-mannanase. (C) 2013 The Institution of Chemical Engineers. Published by Elsevier B.V. All rights reserved.