Evaluation of the production of exopolysaccharides by two strains of the thermophilic bacterium Rhodothermus marinus

AbstractThe thermophile Rhodothermus marinus produces extracellular polysaccharides (EPSs) that forms a distinct cellular capsule. Here, the first data on EPS production in strains DSM4252T and MAT493 are reported and compared. Cultures of both strains, supplemented with either glucose, sucrose, lactose or maltose showed that the EPS were produced both in the exponential and stationary growth phase and that production in the exponential phase was boosted by maltose supplementation, while stationary phase production was boosted by lactose. The latter was higher, resulting in 8.8 (DSM4252T) and 13.7mg EPS/g cell dry weight (MAT493) in cultures in marine broth supplemented with 10g/L lactose. The EPSs were heteropolymeric with an average molecular weight of 8×104Da and different monosaccharides, including arabinose and xylose. FT-IR spectroscopy revealed presence of hydroxyl, carboxyl, N-acetyl, amine, and sulfate ester groups, showing that R. marinus produces unusual sulfated EPS with high arabinose and xylose content

Characterization of high Arabinoxylan oat lines identified from a mutagenized oat population

Oat (Avena sativa) is a nutritionally important cereal crop that is rich in health-promoting dietary fibers, favorable proteins and polar lipids. In this work, ca. 500 random lines of a mutagenized oat population of high genetic variation were screened for arabinoxylan (AX) content. This identified lines with up to 60% higher AX levels in flour from whole seed and up to 100% higher in flour from dehulled seeds, as compared to the original Belinda variety. In addition, the cellular localization of AX was determined in cross-sections of dehulled seeds from three high and one low AX line using a xylan-specific antibody. This revealed variations in the amount and localization of AX between high and low AX lines. The high AX lines will now serve as a starting point in the development of oat varieties with superior health-promoting and rheological properties

HPAEC-PAD analysis for determination of the amino acid profiles in protein fractions from oat flour combined with correction of amino acid loss during hydrolysis

Current derivatization-dependent approaches for amino acid composition analysis of cereal proteins have significant variability due to lack of direct analysis opportunities and loss of amino acids during protein-hydrolysis. To tackle these drawbacks, a novel direct, derivatization-free approach was successfully introduced, using HPAEC-PAD, and applied for analysis of hydrolyzed defatted oat flour and extracted flour protein fractions. The approach ensured reliable detection of amino acids, including L-tryptophan, as well as oxidation products of L-cysteine and L-methionine. A time course study, analysed by nonlinear least-square regression to determine rates of hydrolysis and loss of each amino acid, allowed comparison of the original mass fraction (AA0) of the respective amino acid in the oat flour mixture with the mass fraction obtained after 24 h hydrolysis (AA24). The difference between (AA0) and (AA24) was less than 0.05%, except for L-arginine (0.61%), glycine (0.14%), L-isoleucine (0.27%), and L-tryptophan (0.17%). The (AA0)s obtained corresponded to literature-data, and fitted with the amino acid composition estimated from deduced proteins encoded in the oat genome, except for L-arginine (27%) and L-glutamic acid/L-glutamine (10%). The amino acid composition estimation from sequence data indirectly confirmed that the high presence of L-arginine observed was a result of co-elution with unknown flour components

Macromolecular characterization of high β-glucan oat lines

Oat (Avena sativa) is a cereal grain rich in fibers, proteins, vitamins and minerals. Oats have been linked to several health benefits, such as lowering blood cholesterol levels, counteracting cardiovascular disease and regulating blood sugar levels. This study aimed to characterize two new oat lines with high β-glucan content emanating from ethyl methyl sulphonate mutagenesis on the Lantmännen elite variety Belinda. Two of the mutated lines, and the mother variety Belinda, were profiled for β-glucan, arabinoxylan, total dietary fiber and starch composition. In addition, total lipid and protein content, amino acid composition and β-glucan molecular weights were analyzed. The high levels of β-glucan resulted in a significant increase in total dietary fiber, but no correlation could be established between higher or lower levels of the assayed macromolecules, i.e., between arabinoxylan-, starch-, lipid- or protein levels in the mutated lines compared to the reference. The results indicate separate biosynthetic pathways for β-glucans and other macromolecules and an independent regulation of the different polysaccharides studied. Therefore, ethyl methyl sulphonate mutagenesis can be used to increase levels of multiple macromolecules in the same line.The generous funding provided by ScanOats Industrial Research Center at Lund University made this research possible.Peer reviewe

Valorization of Brewer's spent grain to prebiotic oligosaccharide : Production, xylanase catalyzed hydrolysis, in-vitro evaluation with probiotic strains and in a batch human fecal fermentation model

Brewer's spent grain (BSG) accounts for around 85% of the solid by-products from beer production. BSG was first extracted to obtain water-soluble arabinoxylan (AX). Using subsequent alkali extraction (0.5 M KOH) it was possible to dissolve additional AX. In total, about 57% of the AX in BSG was extracted with the purity of 45–55%. After comparison of nine xylanases, Pentopan mono BG, a GH11 enzyme, was selected for hydrolysis of the extracts to oligosaccharides with minimal formation of monosaccharides. Growth of Bifidobacterium adolescentis (ATCC 15703) was promoted by the enzymatic hydrolysis to arabinoxylooligosaccharides, while Lactobacillus brevis (DSMZ 1264) utilized only unsubstituted xylooligosaccharides. Furthermore, utilization of the hydrolysates by human gut microbiota was also assessed in a batch human fecal fermentation model. Results revealed that the rates of fermentation of the BSG hydrolysates by human gut microbiota were similar to that of commercial prebiotic fructooligosaccharides, while inulin was fermented at a slower rate. In summary, a sustainable process to valorize BSG to functional food ingredients has been proposed

Metabolic engineering of thermophilic bacteria for production of biotechnologically interesting compounds

Many thermophilic bacteria are efficient biomass degraders (producing polysaccharide degrading enzymes and utilizing a great variety of substrates, e.g. lignocellulosic polymers, pentoses, hexoses, as well sugar acids, and sugar alcohols). This makes them interesting organisms as potential cell factories in a circular bioeconomy. Lignocellulosic and marine macroalgal biomasses are regarded as sustainable biorefinery feedstocks for the production of energy carriers and platform and specialty chemicals, thereby meeting impending fossil fuel shortage and counteracting accumulation of greenhouse gasses. However, progress in using thermophilic bacteria that utilize these feedstocks as carbon sources has been hampered by the lack of suitable engineering tools to improve the production profiles of interesting target metabolites as specific synthetic production pathways need to be inserted/modified or existing pathways optimized by metabolic engineering. In this chapter, we review the progress on the use of thermophilic bacteria in metabolic engineering and the available engineering tools and give examples of species for which successful engineering has been accomplished. Today, the majority of thermophilic bacteria targeted for production of compounds of industrial interest by metabolic engineering belong to the phylum Firmicutes (e.g. Thermoanaerobacterium, Caldocellulosiruptor, Geobacillus, and Bacillus), taking advantage of anaerobic catabolic pathways producing organic acids and alcohols. However, there are additional and aerobic species gaining interest concerning biomass degradation and the ability of carbon dioxide fixation as well as production of molecules of interest, and some examples of this are also given

Macromolecular characterization of high β-glucan oat lines

Oat (Avena sativa) is a cereal grain rich in fibers, proteins, vitamins and minerals. Oats have been linked to several health benefits, such as lowering blood cholesterol levels, counteracting cardiovascular disease and regulating blood sugar levels. This study aimed to characterize two new oat lines with high β-glucan content emanating from ethyl methyl sulphonate mutagenesis on the Lantmännen elite variety Belinda. Two of the mutated lines, and the mother variety Belinda, were profiled for β-glucan, arabinoxylan, total dietary fiber and starch composition. In addition, total lipid and protein content, amino acid composition and β-glucan molecular weights were analyzed. The high levels of β-glucan resulted in a significant increase in total dietary fiber, but no correlation could be established between higher or lower levels of the assayed macromolecules, i.e., between arabinoxylan-, starch-, lipid- or protein levels in the mutated lines compared to the reference. The results indicate separate biosynthetic pathways for β-glucans and other macromolecules and an independent regulation of the different polysaccharides studied. Therefore, ethyl methyl sulphonate mutagenesis can be used to increase levels of multiple macromolecules in the same line