In this work, the industrial Saccharomyces cerevisiae PE-2 strain, presenting innate capacity for xylitol accumulation, was engineered for xylitol production by overexpression of the endogenous GRE3 gene and expression of different xylose reductases from Pichia stipitis. The best-performing GRE3-overexpressing strain was capable to produce 148.5 g/L of xylitol from high xylose-containing media, with a 0.95 g/g yield, and maintained close to maximum theoretical yields (0.89 g/g) when tested in non-detoxified corn cob hydrolysates. Furthermore, a successful integrated strategy was developed for the production of xylitol from whole slurry corn cob in a presaccharification and simultaneous saccharification and fermentation process (15% solid loading and 36 FPU) reaching xylitol yield of 0.93 g/g and a productivity of 0.54 g/L·h. This novel approach results in an intensified valorization of lignocellulosic biomass for xylitol production in a fully integrated process and represents an advance towards a circular economy.

The biorefinery concept, consisting in using renewable biomass with economical and energy goals, appeared in response to the ongoing exhaustion of fossil reserves. Bioethanol is the most prominent biofuel and has been considered one of the top chemicals to be obtained from biomass. Saccharomyces cerevisiae, the preferred microorganism for ethanol production, has been the target of extensive genetic modifications to improve the production of this alcohol from renewable biomasses. Additionally, S. cerevisiae strains from harsh industrial environments have been exploited due to their robust traits and improved fermentative capacity. Nevertheless, there is still not an optimized strain capable of turning second generation bioprocesses economically viable. Considering this, and aiming to facilitate and guide the future development of effective S. cerevisiae strains, this work reviews genetic engineering strategies envisioning improvements in 2 nd generation bioethanol production, with special focus in process-related traits, xylose consumption, and consolidated bioprocessing. Altogether, the genetic toolbox described proves S. cerevisiae to be a key microorganism for the establishment of a bioeconomy, not only for the production of lignocellulosic bioethanol, but also having potential as a cell factory platform for overall valorization of renewable biomasses.

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In this work, a sustainable and environmental friendly strategy for the biotechnological production of xylitol was proposed and optimized. For this purpose, corn cob was hydrothermally pretreated at high solid loadings (25%) for an efficient solubilization of xylan in hemicellulose derived compounds, xylooligosaccharides and xylose. Xylose enriched streams were obtained from the enzymatic saccharification of the whole slurry (solid and liquid fraction) resulting from the autohydrolysis pretreatment. The xylitol production in a simultaneous saccharification and fermentation (SSF) process, by the recombinant Saccharomyces cerevisiae PE-2-GRE3 strain, was optimized using different enzyme and substrate (pretreated corn cob solid) loadings by an experimental design. This study demonstrated a significant effect of substrate loading on the production process achieving a maximal concentration of 47 g/L with 6.7 % of pretreated corn cob and 24 FPU/g of enzyme loading, with partial detoxification of the hydrolysate. Furthermore, the 1.42-fold increase in xylitol titer and 1.56-fold increase in productivity achieved in a SSF using an acetic acid free-hydrolysate evidenced the negative effect of acetic acid on the yeast-based xylitol production process. The combination of these green technologies and the optimization of the proposed strategy enhanced the overall xylitol production through the valorization of corn cob.

Cheese whey is the major by-product of the dairy industry, and its disposal constitutes an environmental concern. The production of cheese whey has been increasing, with 190 million tonnes per year being produced nowadays. Therefore, it is emergent to consider different routes for cheese whey utilization. The great nutritional value of cheese whey turns it into an attractive substrate for biotechnological applications. Currently, cheese whey processing includes a protein fractionating step that originates the permeate, a lactose-reach stream further used for valorisation. In the last decades, yeast fermentation has brought several advances to the search for biorefinery alternatives. From the plethora of value-added products that can be obtained from cheese whey, ethanol is the most extensively explored since it is the alternative biofuel most used worldwide. Thus, this review focuses on the different strategies for ethanol production from cheese whey using yeasts as promising biological systems, including its integration in lignocellulosic biorefineries. These valorisation routes encompass the improvement of the fermentation process as well as metabolic engineering techniques for the introduction of heterologous pathways, resorting mainly to Kluyveromyces sp. and Saccharomyces cerevisiae strains. The solutions and challenges of the several strategies will be unveiled and explored in this review.

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