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Biorefinery of biomass-based biofuels and biochemicals by microorganisms is a competitive alternative of traditional petroleum refineries. Zymomonas mobilis is a natural ethanologen with many desirable characteristics, which makes it an ideal industrial microbial biocatalyst for commercial production of desirable bioproducts through metabolic engineering. In this review, we summarize the metabolic engineering progress achieved in Z. mobilis to expand its substrate and product ranges as well as to enhance its robustness against stressful conditions such as inhibitory compounds within the lignocellulosic hydrolysates and slurries. We also discuss a few metabolic engineering strategies that can be applied in Z. mobilis to further develop it as a robust workhorse for economic lignocellulosic bioproducts. In addition, we briefly review the progress of metabolic engineering in Z. mobilis related to the classical synthetic biology cycle of “Design-Build-Test-Learn”, as well as the progress and potential to develop Z. mobilis as a model chassis for biorefinery practices in the synthetic biology era.

Graphical abstract, Highlights • Characterisation of the spent sulphite liquor, a sugar-rich residue was done. • Four chromatographic methods for sugars and derivatives analysis were developed. • Cross-linked Pb+2 columns were suitable for fast and reliable sugars separation. • Cross-linked H+ columns were adequate for acids and furfurals separation. • Methods developed were successfully assayed on woody materials and hydrolysates., Waste from pulp and paper mills consist of sugar-rich fractions comprising hemicellulose derivatives and cellulose by-products. A complete characterisation of the waste streams is necessary to study the possibilities of an existing mill. In this work, four chromatographic methods have been developed to obtain the most suitable chromatographic method conditions for measuring woody feedstocks, lignocellulosic hydrolysates and cellulose pulp in sulphite pulping processes. The analysis of major and minor monosaccharides, aliphatic carboxylic acids and furfurals has been optimised. An important drawback of the spent liquors generated after sulphite pulping is their acidic nature, high viscosity and adhesive properties that interfere in the column lifetime. This work recommends both a CHO-782Pb column for the sugar analysis and an SH-1011 resin-based cross-linked gel column to separate low-molecular-weight chain acids, alcohols and furfurals. Such columns resulted in a good separation with long lifetime, wide pH operating range and low fouling issues.

Furfural, an inhibitor of yeast growth, occurs in xylose-containing lignocellulosic hydrolysates that are potential substrates for the fermentative production of the natural sweetener xylitol from xylose. Effects of furfural on growth and xylitol production by the yeast Candida magnoliae TISTR 5663 are reported. Aerobic as well as oxygen limited conditions were used to assess the effects of furfural. A mathematical model was developed to predict the behavior of this fermentation. The model was validated using independent experimental data. The model and its kinetic parameters are reported. Furfural was found to be a competitive inhibitor of growth. Nevertheless, the presence of a certain amount of furfural (∼0.3 g L −1 ) in the production medium actually improved the productivity of xylitol and its yield from xylose under suitable oxygen limited conditions.

Poacic acid, a decarboxylated product from 8–5-diferulic acid that is commonly found in monocot lignocellulosic hydrolysates, has been identified as a natural antifungal agent against economically significant fungi and oomycete plant pathogens. Starting from commercially available or monocot-derivable ferulic acid, a three-step synthetic procedure has been developed for the production of poacic acid needed for field testing in a controlled agricultural setting. First, ferulic acid was esterified to produce ethyl ferulate in 92% yield. Second, peroxidase-catalyzed free radical dehydrodimerization of ethyl ferulate produced crude diferulates, mainly 8–5-diferulate, in 91% yield. Finally, crystalline poacic acid was obtained in 25% yield via alkaline hydrolysis of the crude diferulates after purification by flash-column chromatography. This new procedure offers two key improvements relevant to large-scale production: 1) bubbling air through the reaction mixture in the second step to remove acetone greatly improves the recovery efficiency of the crude diferulates; and 2) telescoping minor impurities directly into the alkaline hydrolysis step eliminates the need for additional column purifications, thus reducing the overall cost of production and removing a major impediment to process scale-up.

Supplementary data to this article can be found online at https:// doi.org/10.1016/j.biortech.2019.122054., This work presents the exploitation of waste industrial by-products as raw materials for the production of microbial lipids in engineered strains of the filamentous fungus Ashbya gossypii. A lipogenic xylose-utilizing strain was used to apply a metabolic engineering approach aiming at relieving regulatory mechanisms to further increase the biosynthesis of lipids. Three genomic manipulations were applied: the overexpression of a feedback resistant form of the acetyl-CoA carboxylase enzyme; the expression of a truncated form of Mga2, a regulator of the main 9 desaturase gene; and the overexpression of an additional copy of DGA1 that codes for diacylglycerol acyltransferase. The performance of the engineered strain was evaluated in culture media containing mixed formulations of corn-cob hydrolysates, sugarcane molasses or crude glycerol. Our results demonstrate the efficiency of the engineered strains, which were able to accumulate about 40% of cell dry weight (CDW) in lipid content using organic industrial wastes as feedstocks., This work was supported by: the Spanish Ministerio de Economía y Competitividad (grant numbers BIO2014-56930-PandBIO2017-88435R) and the Junta de Castilla y León (grant number SA016P17); the Portuguese Foundation for Science and Technology (FCT) under the scope of the strategic funding of UID/BIO/04469/2019, PhD grant PD/ BD/113812/2015 to R. Silva (Doctoral Program in Applied and Environmental Microbiology), the MultiBiorefinery project (POCI-010145-FEDER-016403) and the BioTecNorte operation (NORTE-010145-FEDER-000004) funded by the European Regional Development Fund under the scope of Norte2020 - Programa Operacional Regional do Norte. DDF was recipient of USAL predoctoral fellowship from the University of Salamanca., info:eu-repo/semantics/publishedVersion