CONICET | Buscador de Institutos y Recursos Humanos

AbstractThe need to reduce waste generation and the use of toxic materials for waste disposal has led to a growinginterest in the development of sustainable bioprocesses using enzymes. Hydrolases can be applied in different fields such as biofuels, pharmaceutical, detergent,oleochemical, organic synthesis andfood industries. Many hydrolases such as, lipases, proteases and glycosyl hydrolases are extracellular, and to make possible its use economically, it is necessary to optimize yields through optimization of culture conditions and/or genetic manipulation to increase the microbial enzyme activities. The objective ofthis work was to optimize hydrolases production, and the design of bioprocess scale-up.The culture medium to obtain high enzyme activities by different strains belongingto Firmicutes phylum was optimized by means of statistical-based designs, followed by apartial purification to assess the enzyme preparations properties. A Box Behnken design-based statistical analysis indicated that the variables that most influenced the lipase andprotease production were peptone and sucrose concentration. Increments on lipase productionup to 33 U/g at 5 g/L of sucrose were observed, while further increments on the carbon sourceconcentration reducedthe enzyme production. The optimized medium (sucrose, 5g/L; peptone, 7.5 g/L; CaCl2, 0.05 M) produced 35.1 U/mL and 4.60 U/mL for lipase andprotease, respectively. Both, protease and lipase, were used to enrich refined fish oils in polyunsaturated fatty acids (ω-3 PUFA); as a result, an increment of 1.2-fold of DHA (docosahexaenoic acid) content was obtained. The examination of the culture media components influencing bacterial growth andendoglucanase production demonstrated that mono and disaccharides are useful substrates forenzyme production. moreover, their combination with carboxymethyl-cellulose (CMC) showed asynergistic effect on enzyme production. Consequently, by using a peptone-based medium amended with sucrose and CMC, a cellulolytic cocktail was efficiently produced in a 1-L stirred tank reactor on batch operation mode, reaching a maximum of 3.12 IU/mL. It is important to point out that the use of simple sugars favors both operative culture conditions and downstream processing. Finally, raw and alkali-pretreated sugarcane bagasse were the carbon sources that better promoted xylanase production. The xylanases obtainedwere further utilized for: (i) a cocktail formulation supplementing commercial enzymesthat improved the glucose releasement (~38 %) from sugarcane pretreated material for secondgeneration ethanol production. (ii) generationof arabino-xylooligosaccharides,potential prebiotics, from the pretreated sugarcane bagasse and wheat bran. Scaling-up these bioprocesses to evaluatethe technical feasibility and the commercial viability is now being performed in a pilot plant with a 150 L Fermentor. Scale parameters were selected for each of the enzymes.