Aspectos do metabolismo de manose em Saccharomyces cerevisiae e produção de etanol a partir da semente de açaí

Abstract

The açaí seed represents about 85% of the fruit’s total mass, generating approximately 1.6 million tons of waste annually, most of which is improperly discarded, causing environmental and sanitary problems. Since around 50% of the seed is composed of mannan, an alternative use is its conversion into mannose followed by fermentation to ethanol. This study aimed to understand the metabolic aspects involved in mannose fermentation and to evaluate ethanol production parameters using hydrolysate from açaí seeds. Metabolic aspects were analyzed in four Saccharomyces cerevisiae laboratory strains with deletions in the genes encoding the enzymes Hxk1, Hxk2, and Glk1, responsible for converting mannose into mannose-6-phosphate, allowing only one enzyme to be expressed per strain. One strain, lacking all three enzymes, was referred to as the triple-deletion strain. All strains grew on mannose in solid medium except for the triple-deletion strain. In liquid medium with 2% sugars, the presence of at least one enzyme was sufficient for mannose consumption and ethanol production, whereas the triple-deletion strain showed no mannose metabolism or ethanol formation. Hxk1 and Hxk2 were more relevant for catabolite repression, showing ethanol production efficiencies of 96.81% and 99.07% (after 6 h), while Glk1 reached 26.54%. Additionally, 13 brewing strains and two industrial fuel-ethanol-producing strains (CAT-1 and Ethanol Red) were evaluated to identify those with higher fermentative potential. Qualitative assays in solid medium showed similar growth in mannose and glucose, and a liquid medium assay with 5% sugars and 1 g/L inoculum led to the selection of CAT-1, ART06, and ART48. Considering the importance of catabolite repression, the repressive effect of mannose was assessed, showing that after 4 h, mannose had a similar effect to glucose in all strains. To study ethanol production, sugar and inoculum concentrations were evaluated. Fermentations with 50 g/L sugars (91% mannose, 9% glucose) and 10 g/L inoculum resulted in sugar consumption of 91.06% (CAT-1), 93.87% (ART06), and 87.49% (ART48), with ethanol production efficiencies of 84.01%, 92.68%, and 96.10%, respectively. Fermentations with 100 g/L sugars reduced efficiencies to 72.11% (CAT-1), 76.75% (ART06), and 72.98% (ART48). Finally, the enzymatic hydrolysate of açaí seed supplemented with nutrients (100 g/L sugars and 10 g/L inoculum) resulted in efficiencies of 84.89% (CAT-1), 81.32% (ART06), and 90.83% (ART48). The study demonstrated that CAT-1, ART06, and ART48 have strong potential for mannose fermentation from açaí seeds, with efficiencies above 85%. The presence of at least one of the enzymes Hxk1, Hxk2, or Glk1 is sufficient for mannose metabolism, with Hxk1 and Hxk2 being crucial for catabolite repression, confirming the technical feasibility of using this agro-industrial residue for second-generation ethanol production.