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Xylitol is a naturally occurring sugar alcohol with a wide range of applications in food, pharmaceutical, and cosmetic industries. It is currently manufactured on a large scale by several companies through the chemical hydrogenation of xylose. The integration of xylose derived from waste stream feedstocks for biotechnological production of xylitol can positively contribute to achieve sustainable development. Wine producing industry annually generates large volumes of by-products and residues that contain high carbohydrate content. In this work, vine pruning residues and grape must were evaluated as feedstocks for xylitol production, using a recombinant Saccharomyces cerevisiae industrial strain [1]. Using grape must as cosubstrate for the bioconversion, the yeast strain directly converted xylose to xylitol, with efficiency similar to reactions that used pure glucose and fructose. Additionally, the capability of the engineered strain to convert D-xylose from pretreated vine pruning residue was also demonstrated. The utilization of both surplus grape must and vine pruning residues, for xylitol production in a multi-feedstock valorization approach, can address the disposal problems of wine producing industry and decrease the overall cost of the process, contributing to economic and environmental sustainability., This study was supported by the Portuguese Foundation for Science and Technology (FCT, Portugal) under the scope of the strategic funding of UIDB/04469/2020, the PhD grant (SFRH/BD/132717/2017 to SLB) and BIOVINO project (0688_BIOVINO_6_E) funded by INTERREG España - Portugal and European Regional., info:eu-repo/semantics/publishedVersion

The utilization of lignocellulosic biomass for the production of biofuels has appeared as a possible solution to alleviate the envisioned depletion of fossil resources. Nevertheless, the attainment of economically viable lignocellulosic-based biorefineries requires the combined production of low value biofuels with high value compounds. 5-hydroxymethylfurfural (HMF) and its derivative 2,5Furandicarboxylic acid (FDCA) have been previously identified as top promising chemicals to be obtained from biomass. Therefore, this work explored the potential of (1) wine residues as renewable sources of hexoses for the production of HMF and (2) Saccharomyces cerevisiae-the preferred microorganism for bioethanol production- as a whole cell biocatalyst for the conversion of HMF into high value derivatives. For that, grape must and pomace were submitted to aqueous microwave treatment at different temperatures and durations for extraction of sugars and their dehydration into HMF. Moreover, different industrial S. cerevisiae strains were compared in terms of HMF conversion, showing different potentials for its oxidation or reduction: with 2,5Hydroxymethylfuran2carboxylic acid (HMFCA) production being favored under more aerobic conditions, while oxygen limitation favored 2,5- Bis(hydroxymethyl)furan (BHMF) accumulation. S. cerevisiae strains were then able to produce these compounds using an HMF-rich medium derived from wine residues. Also, genetic modifications of the yeast resulted in an increase in the accumulation of the further oxidized derivatives FDCA and 5formyl2furancarboxylic acid (FFCA). Taken together, these results prove S. cerevisiae to be a promising whole cell biocatalyst, allowing the envisioning of integrated biorefineries with combined production of biofuels and high value compounds., This study was supported by the Portuguese Foundation for Science and Technology (FCT, Portugal) under the scope of the strategic funding of UIDB/04469/2020, the MIT-Portugal Program (Ph.D. Grant PD/BD/128247/2016 to JTC), and Biovino (0688_BIOVINO_6_E), funded by ERDF through INTERREG España – Portugal., info:eu-repo/semantics/publishedVersion

Additional file 1. Fermentation of ER-X, PE-2-X, CAT-1-X and CA11-X in in corn cob liquor supplemented with 100 g/L glucose at 40 °C; Growth profiles of ER-X, CAT-1-X and CA11-X and corresponding type strains in YPD; Stability evaluation of the integration at the δ-sequences; Fermentation of corn cob liquor with ER-X-2P strain and 2 cycles of cell recycling; Enzymatic activities; Plasmids and primers used for subcloning steps.

info:eu-repo/semantics/publishedVersion

The use of renewable biomass, such as lignocellulosic materials, for the production of biofuels and chemicals within a biorefinery scheme contributes to achieve a sustainable development. Xylitol has been identified as one of the top 12 value added chemicals to be obtained from biomass, and can be produced from hemicellulose-derived xylose through biotechnological processes [1]. In this work, xylitol was produced from xylose (using glucose as co-substrate for co-factor regeneration) in batch fermentations by the industrial Saccharomyces cerevisiae PE-2 strain (over)expressing (1) a wild type xylose reductase from Pichia spititis (XR); (2) a NADH-preferable xylose reductase mutant (mut-XR) from Pichia spititis and (3) the endogenous GRE3 gene which encodes for an unspecific aldose reductase (AR). Maximum yield (0.98 g g-1) was obtained by the strain overexpressing the GRE3 gene. Moreover, the recombinant strain PE-2-GRE3 showed significantly higher xylitol productivity than the laboratory strain, CENPK.113-5D overexpressing the same gene. This strain (PE-2-GRE3) was selected for bioconversion of 160 g L-1 of xylose in a fed-batch fermentation, which resulted in 149 g L-1 of xylitol concentration with a productivity of 1.2 g L-1 h-1. These results open new perspectives and opportunities for the valorisation of hemicellulosic hydrolysates through the production of xylitol within a biorefinery concept., info:eu-repo/semantics/publishedVersion

Microbiotec'17 - Congress of Microbiology and Biotechnology 2017, Background: Lignocellulosic biomass has a recognised potential as a sustainable platform for the production of biofuels and other biochemicals. However, lignin residues and other inhibitory compounds resulting from lignocellulosics pretreatment affect the digestibility of resulting whole slurries. The addition of synergistic proteins that can cooperate with cellulases is an emerging strategy for enhanced lignocellulosics hydrolysis. Carbohydrate-binding modules (CBMs) have been shown to improve the enzymatic hydrolysis of pure cellulose models but their effect on the enzymatic hydrolysis of lignocellulosic materials has not yet been evaluated. Thus, in this work, the potential synergistic effect of a family 3 CBM on the enzymatic saccharification of a pretreated lignocellulosic biomass was studied for the first time. [...], C. Oliveira and D. Gomes acknowledge support from Fundação para a Ciência e a Tecnologia (FCT), Portugal (grants SFRH/BPD/110640/2015 and SFRH/BD/88623/2012, respectively). This study was supported by the FCT under the scope of the GlycoCBMs Project PTDC/AGR-FOR/3090/2012–FCOMP-01-0124-FEDER-027948 and the strategic funding of UID/BIO/04469/2013 unit and COMPETE 2020 (POCI-01-0145-FEDER-006684) and BioTecNorte operation (NORTE-010145-FEDER-000004) funded by the European Regional Development Fund under the scope of Norte2020 - Programa Operacional Regional do Norte., info:eu-repo/semantics/publishedVersion

Food safety and quality are nowadays a major consumers concern. In the dairy industry the fraudulent addition of cheaper/lower-quality milks from nonlegitimate species/breeds compromises the quality and value of the final product. Despite the already existing approaches for identification of the species origin of milk, there is little information regarding differentiation at an intra-species level. In this protocol we describe a low-cost, sensitive, fast, and reliable analytical techniqueRandom Amplified Polymorphic DNA/Sequence Characterized Amplified Region (RAPD/SCAR)capable of an efficient detection of adulterant breeds in milk mixtures used for fraudulent manufacturing of dairy products and suitable for the detection of milk adulteration in processed dairy foods., This study was supported by the Portuguese Foundation for Science and Technology (FCT) under the scope of the strategic funding of UID/BIO/04469/2013 unit and COMPETE 2020 (POCI-01-0145-FEDER-006684), BioTecNorte operation (NORTE-01-0145-FEDER-000004) funded by European Regional Development Fund under the scope of Norte2020— Programa Operacional Regional do Norte, and project “Valor Queijo” (CENTRO-07-0202-FEDER-030372) funded by FCT/ MCTES (PIDDAC) and cofunded by “Fundo Europeu de Desenvolvimento Regional” (FEDER) through “COMPETE— Programa Operacional Factores de Competitividade” (POFC)., info:eu-repo/semantics/publishedVersion

The use of renewable biomass to supply the increasing energetic needs and to partially replace fossil fuels is nowadays recognized as a suitable and desirable alternative to attain a sustainable growth based on a bioeconomy. In spite of the intensive research on lignocellulose-to-ethanol production processes, second generation (2G) bioethanol is still not cost competitive and specific challenges remain. These processes are as a whole substantially more complex than initially thought as several types of biomass may be used as substrate, each with specific challenges. A range of biomass pre-treatments may be applied for biomass fractionation, each with its own specificities and leading to different inhibitor profiles. Finally, different hydrolysis/fermentation schemes may be used. The optimization of these processes has to consider the integration of all the stages of the process and should be done together. One of the key aspects for the development of cost-effective lignocellulose-to-bioethanol processes is the engineering of the yeast strain. There is still a lack of robust and sugars-fast fermentation yeast strains for 2G bioethanol. Recently, we have screened and selected naturally robust yeast strains from industrial environments[1] and engineering some of the more promising strains with the xylose metabolic pathway[2] and inhibitor tolerance genes[3]. We will present the results of the simultaneous engineering of xylose metabolization pathway together with inhibitor tolerance in diverse robust background strains and its performance in different hydrolysates obtained from distinct types of biomass. The heterogeneous outcome of the genetic engineering in different hydrolysates show that tolerance and xylose engineering must be customized to the strain background and hydrolysate used in the process. The results obtained highlight that yeast development must not only be integrated in the process but it must also be tailor-made for each specific process. [1]Pereira et al.(2014)Bioresource Technology 161:192-199. [2]Romani et al.(2015)Bioresource Technology 179:150-158. [3]Cunha et al.(2015)Bioresource Technology 191:7-16.