Use of agro waste biomass for alpha-amylase production by Anoxybacillus amylolyticus. Purification and properties

Knowledge accumulated from fundamental and applied studies on Anoxybacillus suggests that this genus can serve as a good alternative in many applications related to starch and lignocellulosic biomasses, waste treatment, enzyme technology, and bioenergy production. We investigated the purification, biochemical characterization and immobilization of a thermostable ?-amylase from the thermophilic Anoxybacillus amylolyticus, strain MR3CT, isolated in Antarctica and its production on vegetable wastes. In particular, the rhizome from Arundo donax, waste biomass of Cynara cardunculus and potato peels were tested either in Submerged Fermentation (SmF) and Solid State Fermentation (SSF) conditions. The amylase from A. amylolyticus, with a molecular weight of about 60 kDa, displayed an optimum enzyme activity at 60°C and pH 5.6. Moreover, by retaining up to 70% of total activity after 48 h at 60°C, it showed high thermostability in the presence of 2 mM calcium ion. The immobilized enzyme maintained the 48% of its initial activity after the sixth reuse. The optimal conditions for its production in SmF were achieved at 60°C for 24 h with 1% of rhizome from Arundo donax, which was about 2126 U/gds. SSF cultures reached maximum ?-amylase yield (102 U/gds) when grown on waste biomass of Cynara cardunculus as substrate, with a substrate-water ratio of 1:1 (w/v), and incubation at 60°C for 4 days. In this study, rhizome of A. donax resulted to be a good substrate for amylase production in SmF thus allowing a cheaper alternative to obtain amylolytic enzymes. Indeed by using rhizomes from A. donax as growth substrate it was possible to recovery an amylase activity level higher than that obtained by synthetic complex medium. Amylase production was also investigated under SSF conditions by using the above listed wastes as sole carbon source for A. amylolyticus growth. Under these conditions, C. cardunculus gave a higher enzyme yield per reactor volume.