Purification of CBS 819.72 α-amylase by aqueous two-phase systems: Modelling using Response Surface Methodology

The partition and purification of α-amylase from a culture supernatant of Aspergillus oryzae CBS 819.72 was made in aqueous two-phase system (ATPS). According to bibliography and preliminary studies, the factors polyethylene glycol (PEG) molecular weight (MPEG) and concentration (CPEG), buffer type (BU) and concentration (CBU), temperature (T), salt nature (SALT) and concentration (CSALT), bioligand (BL) and concentration (CBL) and pH were investigated using a Plackett–Burman design to identify the factors affecting separation. Taking into consideration a simultaneous increase in enzyme recovery (RY) and purification factor (PF), the best performance of the system was obtained at 4 °C and pH 6 using PEG 8000 g/mol, citrate buffer, KCl and sucrose. Experimental Box–Behnken design together with the Response Surface Methodology (RSM) have been used to find optimum CPEG, CCitrate and CSALT. Quadratic models were predicted for PF and RY in the top phase and a better compromise between these two parameters can be found by superimposing the contour plots of PF and RY for 8% citrate. A region in the experimental space can be defined where the purification factor is always higher than 3 with yields exceeding 65%.