Alpha-amylase is one of the most employed enzymes in the food industry because of its capacity to degrade starch, improving the organoleptic and nutritional properties of food products. Thus, it is very important to develop novel industrial production processes for its production. A submerged fermentation process using Aspergillus oryzae was optimised to modulate the most important factors affecting alpha-amylase production. Flour mill waste, an abundant worldwide agro-industrial residue, was used as substrate due to its starch-rich composition. This residue was able to avoid catabolite repression during the fermentation, acting as a slow-release substrate. The process optimization led to a maximum yield of alpha-amylase production of 14076 ± 2346 U/L. The secretome of the fungi in the tested conditions was analysed by LC-MS, showing that two isoforms of alpha-amylase (amy-1 and amy-3) were produced. Finally, a two-step process was developed to purify alpha-amylase, consisting of fractional precipitation using (NH4)2SO4 followed by hydrophobic interaction chromatography. The purification allowed a four-time concentration of alpha-amylase, with an eleven-time purification factor and 65 % recovery and a reduction of the proteolytic activity to 14%.
