Over 8 million tonnes of sugar beet are grown annually in the UK. Sugar beet pulp (SBP) is the main by-product of sugar beet processing which is currently dried and sold as a low value animal feed. SBP is a rich source of carbohydrates, mainly in the form of cellulose and pectin, including d-glucose (Glu), l-arabinose (Ara) and d-galacturonic acid (GalAc). This work describes the technical feasibility of an integrated biorefinery concept for the fractionation of SBP and conversion of these monosaccharides into value-added products. SBP fractionation is initially carried out by steam explosion under mild conditions to yield soluble pectin and insoluble cellulose fractions. The cellulose is readily hydrolysed by cellulases to release Glu that can then be fermented by a commercial yeast strain to produce bioethanol at a high yield. The pectin fraction can be either fully hydrolysed, using physico-chemical methods, or selectively hydrolysed, using cloned arabinases and galacturonases, to yield Ara-rich and GalAc-rich streams. These monomers can be separated using either Centrifugal Partition Chromatography (CPC) or ultrafiltration into streams suitable for subsequent enzymatic upgrading. Building on our previous experience with transketolase (TK) and transaminase (TAm) enzymes, the conversion of Ara and GalAc into higher value products was explored. In particular the conversion of Ara into l-gluco-heptulose (GluHep), that has potential therapeutic applications in hypoglycaemia and cancer, using a mutant TK is described. Preliminary studies with TAm also suggest GluHep can be selectively aminated to the corresponding chiral aminopolyol. The current work is addressing the upgrading of the remaining SBP monomer, GalAc, and the modelling of the biorefinery concept to enable economic and Life Cycle Analysis (LCA).
The search for lead product with beneficial pharmacological properties has become a great challenge and costly. Extraction and synthetic modification of bioactive compounds from natural resources has gained great attention and is cost effective. In this study, kojic acid was produced from fungal fermentation, using sago waste as substrate, and chemically incorporated with chalcones and azobenzene to form a series of kojic ester derivatives and evaluated for antibacterial activities. Kojic ester bearing halogenated chalcone demonstrated active inhibition against Staphylococcus aureus compared to that of standard ampicillin. The inhibition increased as the electronegativity of halogens decreased, while incorporation of azobenzene derivatives on kojic acid backbone demonstrated fair antibacterial activity against Escherichia coli with minimum inhibitory concentration (MIC) of 190–330 ppm. The presence of C=C and N=N reactive moieties in both chalcone and azo molecules contributed to the potential biological activities of the kojic acid ester.
BACKGROUND: This work explores the feasibility of vinasse as an inexpensive feedstock for industrial biocatalyst production within the context of an integrated sugar beet biorefinery. As an exemplar, production of CV2025 -Transaminase ( -TAm) in Escherichia coli BL21 was studied. RESULTS:Characterisation of vinasse showed that it comprised mainly of glycerol along with several reducing sugars, sugar alcohols, acetate, polyphenols and protein. Preliminary results showed E. coli BL21 cell growth and CV2025 -TAm production were feasible in cultures using 17% to 25% (v/v) vinasse with higher concentrations demonstrating inhibitory effects. The D-galactose present in vinasse facilitated auto-induction of the pQR801 plasmid enabling CV2025 -TAm expression without addition of expensive Isopropyl--D-thiogalactopyranoside (IPTG). Assessment of different vinasse pre-processing options confirmed simple dilution of the vinasse was sufficient to reduce the concentration of polyphenols to below inhibitory levels. Optimisation experiments, carried out using a controlled, 24-well microbioreactor platform, showed supplementation of diluted vinasse medium with 10 g L −1 yeast extract enabled enhancements of 2.8, 2.5, 5.4 and 3-fold in specific growth rate, maximum biomass concentration, CV2025 -TAm volumetric and specific activity, respectively. Investigation into the metabolic preferences of E. coli BL21 when grown in vinasse showed a preference for D-mannitol utilisation before simultaneous metabolism of glycerol, D-xylitol, D-dulcitol and acetate. Scale-up of optimised conditions for batch CV2025 -TAm production to a 7.5 L stirred tank reactor (STR) was demonstrated based on matched volumetric mass transfer coefficient (k L a). The results showed good comparability with respect to cell growth, substrate consumption and CV2025 -TAm production representing over a 700-fold volumetric scale translation. Further enhancements in CV2025 -TAm production were possible in the STR when operated at higher k L a values. CONCLUSION: This work describes the promising application of vinasse for production of microbial enzymes and insights into carbon source utilisation in complex feedstocks. Exploitation of vinasse as a fermentation feedstock could be further extended to other processes involving different microorganisms and target enzymes.
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