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Simple Summary: This study is the first to elucidate the milk oligosaccharide profile of globally distributed cattle breeds such as Holstein and Ayrshire, which are widely utilized in the dairy industry, with the endangered Northern Finncattle breed, native to Lapland, Finland. Oligosaccharides are sugars with complicated structures that play crucial roles in neonatal development and gut health. We investigated the variations in neutral, sialylated, and fucosylated oligosaccharides among these breeds. Although similar oligosaccharide distributions were observed in all breeds, Ayrshire milk was shown to contain oligosaccharides with more intricate structures, whereas Northern Finncattle and Holstein milk contained a higher level of sialylated oligosaccharides, which are known for their role in promoting brain development. Holstein had the lowest abundance of fucosylated oligosaccharides among the three breeds. These findings enhance our understanding of the milk composition of cattle breeds, which can inform breeding programs and dairy product development, ultimately contributing to improved nutritional and health outcomes in society. In addition, these findings indicate significantly different OS content in NF, which may inspire further preservation efforts based on enhancing genetic diversity. Milk oligosaccharides are complex carbohydrates composed of various monosaccharide units linked together by glycosidic bonds. They play an essential role in promoting gut health by fostering beneficial bacteria, supporting the development of the immune system, and protecting against infections and diseases. This work compared the oligosaccharide profiles in widely utilized breeds such as Holstein and Ayrshire (Nordic Red), with the native Northern Finncattle, which is considered an endangered breed. Oligosaccharides were extracted from milk and analyzed by liquid chromatography–mass spectrometry. The composition and relative abundance of the identified oligosaccharides were characterized and compared. The statistical analyses showed that neutral, sialylated, and fucosylated oligosaccharides vary among the breeds. Ayrshire and Northern Finncattle oligosaccharides formed a cluster, while Holstein's profile shared features with both Ayrshire and Northern Finncattle. Holstein had the lowest abundance of fucosylated OS among the three breeds, with Ayrshire having the highest content followed by Northern Finncattle. The relatively higher sialylated over neutral content of Northern Finncattle is an important feature that should be preserved. Ayrshire is a good candidate to recover more diverse oligosaccharides with potential gut health implications for consumers. [ABSTRACT FROM AUTHOR]

β-Amylase (α-1,4-glucan maltohydrolase; EC 3.2.1.2) is known to be an important enzyme in beer brewing. To identify the contribution of β-amylase in more detail, two β-amylase-deficient barley, 'Backcrossed line (BCL)-Haruna' and 'Nittakei 81', were used for the investigation of malt quality and brewing characteristics. 'BCL-Haruna' was bred by the backcross method for the introduction of the β-amylase-deficient trait from landrace barley ('Tibet Violet 1') to Japanese traditional malting barley 'Haruna Nijo'. 'Nittakei 81' was bred by crossing between Japanese malting barley 'Sukai Golden' and 'Tibet Violet 1'. With regards to the malting quality under 41.0% target cast moisture in micro-malting, the diastatic power (DP) and apparent attenuation limit (AAL) of 'BCL-Haruna' and 'Nittakei 81' were 408 oWK and 416 oWK and 12.4% and 14.8% lower than those of the control variety 'Haruna Nijo'. Oligosaccharide analysis indicated that the maltose concentration of the 'Nittakei 81' wort was 19.7 g/L lower in comparison to a standard European malting barley 'Propino' wort, although the maltotetraose and maltopentaose concentrations of the 'Nittakei 81' wort were 5.6 g/L and 2.7 g/L higher than those of the 'Propino' wort. After fermentation, the alcohol concentration of the 'Nittakei 81' beer was 0.67% lower compared with the 'Propino' beer, and the extract and concentrations of maltotetraose and maltopentaose of the 'Nittakei 81' beer were 5.0 g/L and 2.5 g/L higher than those of the 'Propino' beer. The results indicate that the β-amylase-deficient barley has a unique malting quality, especially lower DP and AAL, which might induce inadequate starch degradation or fermentation. However, this trait caused no serious issues during the brewing process, suggesting the possibility of β-amylase-deficient barley as a unique brewing material including low-alcoholic and non-alcoholic beer production with a specific saccharide composition. [ABSTRACT FROM AUTHOR]

Carboxymethylglucans (CMGs) are widely used semisynthetic polyelectrolytes, e.g., for pharmaceuticals. They are produced in heterogeneous processes on activated starch granules or cellulose fibers. In contrast to neutral ether derivatives, a lower DS in the range, commonly between 0.6 and 1.2, is sufficient to achieve the water solubility of CM cellulose. The high proportion of unsubstituted domains, which could aggregate and therefore only swell and form gel particles but do not dissolve, places higher demands on the statistical distribution of the substituents. The knowledge of regioselectivity, essential for the interpretation of higher structural-level data, can be obtained by various methods, preferentially by CE/UV after hydrolysis. To study the distribution of substituents at the polymer level by mass spectrometric (MS) analysis, partial random depolymerization is required. Due to the ionic character and acid functionality, all the attempts of the direct depolymerization of CMG and further sample preparation suffered from bias, side reactions, and multiple ion formation in MS. Finally, the transformation of CMGs to the corresponding hydroxyethylglucans (HEGs) by the reduction of the esterified carboxy groups with LiAlH4 opened the window for quantitative oligomer MS analysis. While the CM amyloses were reduced quantitatively, the transformation of the CMC was only about 60% but without the formation of mixed CM/HE ethers. [ABSTRACT FROM AUTHOR]

AbstractChitosan oligosaccharides are biopolymers with a wide range of potential applications in various fields. This biopolymer is diverse and promising, and current research is investigating its capabilities for improved drug delivery. As chitosan oligosaccharide has the potential to be used as a drug delivery option, the purpose of this study was to examine its physicochemical characteristics and its potential for drug delivery. In this study, the pharmacokinetic properties of chitosan oligosaccharide were studied through Insilco investigation, which revealed that it is an extremely soluble and effective drug delivery candidate because it does not inhibit CYP isoenzymes and has a log Kp of −12.10 cm/s. It belongs to toxicity class 6 for acute oral toxicity, with an average similarity of 87.5% and a prediction accuracy of 70.97%. Additionally, XRD peak analysis revealed that the material was amorphous, as the peak appeared at 2θ = 24.62°, indicating the absence of well-defined crystalline areas. This characteristic makes the material more suitable for customization in many applications such as drug delivery and tissue engineering. FTIR, SEM, and TGA analysis were performed to gain a better understanding. These findings also emphasize the distinctive qualities and benefits of the oligosaccharides in this domain. Application of chitosan oligosaccharides in the development of efficient drug delivery systems. In the future, it would be more effective, targeted, and safe, with potent therapeutic efficacy for drug delivery. [ABSTRACT FROM AUTHOR]

Understanding the relation between enzyme domain structure and catalytic activity is crucial for optimal engineering of novel enzymes for lignocellulose bioconversion. Xylanases with varying specificities are commonly used to valorise the hemicellulose arabinoxylan (AX), yet characterization of specific arabinoxylanases remain limited. Two homologous GH5_34 arabinoxylanases, Hh Xyn5A and Ct Xyn5A, in which the two domains are connected by a 40-residue linker, exhibit distinct activity on AX, yielding different reaction product patterns, despite high sequence identity, conserved active sites and similar domain composition. In this study, the carbohydrate binding module 6 (CBM6), or the inter domain linker together with CBM6, were swapped to investigate their influence on hydrolytic activity and oligosaccharide product pattern on cereal AXs. The variants, with only CBM6 swapped, displayed reduced activity on commercial wheat and rye AX, as well as on extracted oat fibre, compared to the original enzymes. Additionally, exchange of both linker and CBM6 resulted in a reduced ratio of enzyme produced in soluble form in Escherichia coli cultivations, causing loss of activity of both Hh Xyn5A and Ct Xyn5A variants. Analysis of oligosaccharide product patterns applying HPAEC–PAD revealed a decreased number of reaction products for Ct Xyn5A with swapped CBM6, which resembled the product pattern of Hh Xyn5A. These findings emphasize the importance of the CBM6 interactions with the linker and the catalytic domain for enzyme activity and specificity, and underlines the role of the linker in enzyme structure organisation and product formation, where alterations in linker interactions with the catalytic and/or CBM6 domains, influence enzyme-substrate association and specificity. [ABSTRACT FROM AUTHOR]

Phosphoesterification is the only naturally occurring covalent starch modification identified to date, and it has a major impact on overall starch metabolism. The incorporation of phosphate groups mediated by dikinases [α-glucan, water dikinase (GWD), EC 2.7.9.4; phosphoglucan, water dikinase (PWD), EC 2.7.9.5] massively alters the starch granule properties; however, previous studies did not determine whether the starch-related dikinases bind the phosphate to the glucosyl units within the amylopectin molecules in a specific pattern or randomly. In order to answer this challenging question, a number of approaches were initially pursued until a protocol could be established that enabled a massive step forward in the in vitro analysis of phosphorylated glucan chains obtained from starch. For this purpose, phosphorylation by GWD was investigated, including the final state of phosphorylation i.e., the state of substrate saturation when GWD lacks further free hydroxyl groups at OH-C6 for the catalysis of monophosphate esters. Since the separated phosphorylated glucan chains were required for the analysis, isoamylase digestion was performed to cleave the α-1,6-glycosidic bonds and to allow for the removal of the huge number of existing neutral chains by means of anion exchange chromatography. Via Matrix-Assisted Laser Desorption/Ionization–Time of Flight (MALDI-TOF) MS and MALDI-MS/MS, the phosphorylated α-glucan chains were analysed, and the position of the phosphate group within the chain in relation to the reducing end was determined. Here, we demonstrate a protocol that enables the analysis of phosphorylated oligosaccharides, even in small quantities. [ABSTRACT FROM AUTHOR]

Alginate is a commercially important polysaccharide composed of mannuronic acid and its C5 differential isomer guluronic acid. Comprehensive research on alginate and alginate lyases requires efficient and precise analytical methods for alginate oligosaccharides. In this research, high-performance anion exchange chromatography (HPAEC) in parallel with pulsed amperometric detection (PAD) and mass spectrometry (MS) was applied to the analysis of oligosaccharides obtained by alginate lyase. By optimizing the chromatographic conditions including mobile phase concentration, flow rate, and elution gradient, the analysis of a single sample could be completed in 30 min. Seven unsaturated alginate oligosaccharides were separated and identified through their analysis time observed with PAD, including all structurally different unsaturated disaccharides and trisaccharides. The quantitative analysis of seven oligosaccharides was performed based on the quantitative capability of PAD. The method exhibited adequate linearity and precision parameters. All the calibration curves showed good linearity at least in the concentration range of 0.002 to 0.1 mg/mL. The HPAEC-PAD/MS method provides a general and efficient online method to analyze alginate oligosaccharides. [ABSTRACT FROM AUTHOR]

In the crucial early months of life, an infant's health, growth, and safeguarding are paramount. Breastfeeding stands out as the optimal and natural means of nourishment, offering profound benefits for their development and overall wellness. Within breast milk, a rich array of bioactive components, including hormones, oligosaccharides, and immunoglobulins, provide neonates with an ideal nutritional balance. Among these, Human Milk Oligosaccharides (HMOs) rank prominently, serving both as prebiotics and shields against various neonatal ailments. Particularly, compounds like 2'-Fucosyllactose (2'-FL) and Lacto-N-Neotetraose (LNnT) have been deemed safe for integration into infant formulas, amplifying their benefits. Extensive research underscores the manifold advantages of HMOs, from modulating gut microbiota to bolstering immune function and thwarting pathogenic invaders. HMOs also exhibit a nuanced interplay with maternal genetics, influencing their quantity and diversity. Infants nursed by secretor mothers tend to enjoy enhanced HMO benefits compared to non-secretor counterparts. Incorporating HMOs like Lacto-N-Neotetraose and 2'-Fucosyllactose into infant formulas represents a pivotal stride in optimizing child nutrition. This review offers a thorough exploration of recent research on HMOs, delving into their varied types, concentrations, and compositions, while emphasizing their profound impact on infant health and safety. By consolidating current research findings and recent progress, this study seeks to clarify the diverse effects of HMOs on different facets of human physiology, immune system maturation, modulation of gut microbiota, neurological functioning, and lifelong health implications. Emphasizing the pivotal role of HMOs in influencing human well-being from infancy to maturity, this review emphasize the necessity for continued research and potential utilization in clinical settings and nutritional science. [ABSTRACT FROM AUTHOR]

Pequi is a native and popular fruit in Cerrado biome. The internal yellow-orange mesocarp is the edible fraction of the fruit, but its shell (peel and external mesocarp), which comprises 80% of the fruit, is not used by the agro-industry during fruit processing. There is a growing interest in the reduction of food loss and waste because of environmental, economic, and social impacts. So this study evaluated the chemical composition, antioxidant capacity, and in vitro prebiotic activity of pequi shell flour. Pequi shell flour was obtained from the lyophilization and milling of pequi shell. The content of dietary fibers, oligosaccharides, sugars, organic acids, total phenolics and tannins, polyphenol profile, and antioxidant capacity was determined in pequi shell flour. In addition, its prebiotic activity was evaluated on growth and metabolism of probiotics Lactobacillus and Bifidobacterium strains. Pequi shell flour has a high content of dietary fibers (47.92 g/100 g), soluble fibers (18.65 g/100 g), raffinose (2.39 g/100 g), and phenolic compounds (14,062.40 mg gallic acid equivalents/100 g). For the first time, the polyphenols epigallocatechin gallate, epicatechin, and procyanidin B2 were identified in this by-product. Pequi shell flour promoted greater growth of Lacticaseibacillus casei L-26 (at 24–48 h) and Bifidobacterium animalis subsp. lactis BB-12, as well as higher prebiotic activity scores than fructooligosaccharides (standard prebiotic). Pequi shell flour is rich in prebiotic compounds and has a high antioxidant and prebiotic potential. The promising results encourage its use as an ingredient with antioxidant and potential prebiotic properties to elaborate new functional foods and nutraceuticals. [ABSTRACT FROM AUTHOR]

The sample preparation including labeling and clean‐up represents a key analytical step in the analysis of oligosaccharides and glycans by either chromatographic or electrophoretic separation methods. Although the majority of labeling has been performed by neutral and/or negatively charged tags, the introduction of a positive charge into the saccharide molecule can significantly improve the analysis, especially with mass spectrometry detection. In this work, we present the evaluation of five solid‐phase extraction sorbents differing in extraction chemistry for the clean‐up and concentration of positively labeled maltooligosaccharides from the reaction mixtures. Maltooligosaccharides containing four to seven glucose units were labeled by cationic tags (2‐aminoethyl)trimethylammonium chloride and (carboxymethyl)trimethylammonium chloride hydrazide and the extraction conditions were optimized followed by electrophoretic analysis with conductivity detection. The effects of the solid‐phase extraction sorbent chemistry, extraction conditions, and sample composition are discussed. All tested sorbents were capable of cleaning up maltooligosaccharides from the reaction mixtures to some extent after optimization of the solid‐phase extraction procedure (51.9%–98.9% recovery). The best‐rated amide‐based sorbent was used to process the sample of N‐linked glycans enzymatically released from ribonuclease B. [ABSTRACT FROM AUTHOR]

Sweet corn is frequently consumed in the US and contains carbohydrates as major macronutrients. This study examined the effects of blanching, freezing, and canning on carbohydrates in sweet corn. Fresh bi-color sweet corn was picked in the field and processed immediately into frozen and canned samples. Simple sugars, starch, and dietary fiber (DF) (including total DF (TDF), insoluble DF (IDF) and two fractions of soluble DF (SDF)) were measured according to the AOAC methods. Additional glycomic analysis including oligosaccharides, monosaccharide composition of total polysaccharides (MCTP) and glycosidic linkage of total polysaccharides (GLTP) were analyzed using UHPLC-MS. Sucrose is the major simple sugar, and IDF is the main contributor to TDF. Sucrose and total simple sugar concentrations were not altered after blanching or freezing but were significantly reduced in canned samples. Kestose was the only oligosaccharide identified in sweet corn and decreased in all heat-treated or frozen samples. Starch content decreased in frozen samples but increased in canned samples. While two SDF fractions did not differ across all samples, blanching, freezing and canning resulted in increases in TDF and IDF. Six monosaccharides were identified as major building blocks of the total polysaccharides from MCTP analysis. Glucose and total monosaccharide concentrations increased in two canned samples. GLTP was also profoundly altered by different food processing methods. This study provided insights into the changes in the content and quality of carbohydrates in sweet corn after food processing. The data are important for accurate assessment of the carbohydrate intake from different sweet corn products. [ABSTRACT FROM AUTHOR]

Tamm–Horsfall protein (THP) is a highly N-glycosylated protein from epithelial cells of the ascending limb of Henle loop. It is secreted into the urine as part of the innate immune response against uropathogenic pathogens. As women are more likely to suffer from urinary tract infections, biomedical studies were conducted to investigate sex-differences in THP excretion, as well as differences in the THP N-glycosylation pattern. A total of 238 volunteers (92 men, 146 women, 69 with hormonal contraceptives) participated in this study, providing urine samples. Women showed a clear tendency to have higher THP concentration and excretion rates than men (p < 0.16). Regular intake of hormonal contraceptives had no significant influence on urinary THP concentration compared to no regular intake. The individual N-glycosylation pattern of THP in urine samples from randomly selected individuals (10 female, 10 male) was investigated after enzymatic release and MS analysis of the oligosaccharides. Female subjects tended to have an increased proportion of oligomannose type N-glycans and non-fucosylated glycans, whereas men had an increased proportion of fucosylated complex-type glycans. The higher level of oligomannose-type glycans in THP from women might be explained by a self-defence mechanism to overcome the higher infections pressure by the female anatomical properties. [ABSTRACT FROM AUTHOR]

Carbohydrates play essential roles in a variety of biological processes that are dictated by their structures. However, characterization of carbohydrate structures remains extremely difficult and generally unsolved. In this work, a de novo mass spectrometry-based workflow was developed to isolate and structurally elucidate oligosaccharides to provide sequence, monosaccharide compositions, and glycosidic linkage positions. The approach employs liquid chromatography-tandem mass spectrometry (LC-MS/MS)-based methods in a 3-dimensional concept: one high performance liquid chromatography-quadrupole time-of-flight mass spectrometry (HPLC-QTOF MS) analysis for oligosaccharide sequencing and two ultra high performance liquid chromatography-triple quadrupole mass spectrometry (UHPLC-QqQ MS) analyses on fractionated oligosaccharides to determine their monosaccharides and linkages compositions. The workflow was validated by applying the procedure to maltooligosaccharide standards. The approach was then used to determine the structures of oligosaccharides derived from polysaccharide standards and whole food products. The integrated LC-MS workflow will reveal the in-depth structures of oligosaccharides.

Background: Salt stress is one of the key factors limiting rice production. Alginate oligosaccharides (AOS) enhance plant stress resistance. However, the molecular mechanism underlying salt tolerance in rice induced by AOS remains unclear. FL478, which is a salt-tolerant indica recombinant inbred line and IR29, a salt-sensitive rice cultivar, were used to comprehensively analyze the effects of AOS sprayed on leaves in terms of transcriptomic and metabolite profiles of rice seedlings under salt stress. Results: In this experiment, exogenous application of AOS increased SOD, CAT and APX activities, as well as GSH and ASA levels to reduce the damage to leaf membrane, increased rice stem diameter, the number of root tips, aboveground and subterranean biomass, and improved rice salt tolerance. Comparative transcriptomic analyses showed that the regulation of AOS combined with salt treatment induced the differential expression of 305 and 1030 genes in FL478 and IR29. The expressed genes enriched in KEGG pathway analysis were associated with antioxidant levels, photosynthesis, cell wall synthesis, and signal transduction. The genes associated with light-trapping proteins and RLCK receptor cytoplasmic kinases, including CBA, LHCB, and Lhcp genes, were fregulated in response to salt stress. Treatment with AOS combined with salt induced the differential expression of 22 and 50 metabolites in FL478 and IR29. These metabolites were mainly related to the metabolism of amino and nucleotide sugars, tryptophan, histidine, and β -alanine. The abundance of metabolites associated with antioxidant activity, such as 6-hydroxymelatonin, wedelolactone and L-histidine increased significantly. Combined transcriptomic and metabolomic analyses revealed that dehydroascorbic acid in the glutathione and ascorbic acid cycles plays a vital role in salt tolerance mediated by AOS. Conclusion: AOS activate signal transduction, regulate photosynthesis, cell wall formation, and multiple antioxidant pathways in response to salt stress. This study provides a molecular basis for the alleviation of salt stress-induced damage by AOS in rice. [ABSTRACT FROM AUTHOR]

This study aims to investigate the concentration of 2'-FL HMO and its relationship with infant health status, then examine their correlation to macroscopic stool examination. A case-control study was conducted among 103 mother-infant pairs in 3 primary health care in Malang City, Indonesia. HPLC analyzed HMO quantification and fecal assessment by gross macroscopic stool examination. The findings showed that 49 infants had stunted nutrition status, and 54 had not stunted nutritional status. Among the group of stunted infants came from mothers with secretor-positive status (40.81%), while all infants with not stunted nutritional status came from mothers with secretor-positive status (100%). However, the status of secretor mothers to nutritional status was not significantly related (p>0.05). Levels of 2'-FL HMO in breast milk in stunted infants had a lower average than in non-stunted infants (1.21 mg/L vs. 1.40 mg/L). After analysis with the Mann-Whitney Test, 2'-FL HMO levels had a significant relationship with the baby's nutritional status, the yellow color in infant stool, and mucus in large amounts of stool (p>0.05). 2'-FL HMO has a significant role in the nutritional status of infants. Further analysis is needed to validate the macroscopic assessment of stool to detect inflammation and indigestion in stunted infants. [ABSTRACT FROM AUTHOR]

Fucoidans are discussed as antiviral agents, and fucoidan from Undaria pinnatifida (UpF), in particular has gained interest as potential food additive in antinoroviral strategies. As the competitive blocking activity of antinoroviral agents increases with the valency of terminal nonreducing fucose on the competitor, an effective processing of fucoidans to inhibitory oligosaccharides will depend on basic structural features of the polysaccharide. We demonstrate increased antiviral binding activity of processed low-mass UpF generated by hydrothermal degradation contrasting with decreased efficacy of low-mass fucoidan from Fucus vesiculosus. As this finding is in conflict with current structural models of UpF, we undertook a re-investigation of the glycan backbone in UpF. Applying solvolytical desulfation combined with enzymatic cleavage of low-mass fucoidan by endo-β6-galactanase and terminal labeling of oligosaccharides by deutero-reduction and bis-5-phenyl-3-methyl-1-pyrazolone (PMP) substitution, evidence from mass spectrometry and methylation linkage analysis of the oligosaccharides indicates that fucoses and galactoses in the glycan backbone are organized in homomeric blocks, where oligo-fucoses branch off from a galactane-type core: Fuc(1-3Fuc)n1-3[Gal(1-6Gal)n1-6]Gal(1-6Gal)n. [ABSTRACT FROM AUTHOR]

Intestinal mucous layers mediate symbiosis and dysbiosis of host-microbe interactions. These interactions are influenced by the mucin O-glycan degrading ability of several gut microbes. The identities and prevalence of many glycoside hydrolases (GHs) involved in microbial mucin O-glycan breakdown have been previously reported; however, the exact mechanisms and extent to which these GHs are dedicated to mucin O-glycan degradation pathways warrant further research. Here, using Bifidobacterium bifidum as a model mucinolytic bacterium, we revealed that two ß-N-acetylglucosaminidases belonging to the GH20 (BbhI) and GH84 (BbhIV) families play important roles in mucin O-glycan degradation. Using substrate specificity analysis of natural oligosaccharides and O-glycomic analysis of porcine gastric mucin (PGM) incubated with purified enzymes or B. bifidum carrying bbhI and/or bbhIV mutations, we showed that BbhI and BbhIV are highly specific for ß-(1→3)- and ß-(1→6)-GlcNAc linkages of mucin core structures, respectively. Interestingly, we found that efficient hydrolysis of the ß-(1→3)-linkage by BbhI of the mucin core 4 structure [GlcNAcß1-3(GlcNAcß1-6)GalNAca-OThr] required prior removal of the ß-(1→6)-GlcNAc linkage by BbhIV. Consistent with this, inactivation of bbhIV markedly decreased the ability of B. bifidum to release GlcNAc from PGM. When combined with a bbhI mutation, we observed that the growth of the strain on PGM was reduced. Finally, phylogenetic analysis suggests that GH84 members may have gained diversified functions through microbe-microbe and host-microbe horizontal gene transfer events. Taken together, these data strongly suggest the involvement of GH84 family members in host glycan breakdown. [ABSTRACT FROM AUTHOR]

Crimean-Congo haemorrhagic fever virus (CCHFV) is a pathogen of increasing public health concern, being a widely distributed arbovirus and the causative agent of the potentially fatal Crimean-Congo haemorrhagic fever. Hazara virus (HAZV) is a genetically and serologically related virus that has been proposed as a surrogate for antiviral and vaccine testing for CCHFV. Glycosylation analysis of HAZV has been limited; first, we confirmed for the first time the occupation of two N-glycosylation sites in the HAZV glycoprotein. Despite this, there was no apparent antiviral efficacy of a panel of iminosugars against HAZV, as determined by quantification of the total secretion and infectious virus titres produced following infection of SW13 and Vero cells. This lack of efficacy was not due to an inability of deoxynojirimycin (DNJ)-derivative iminosugars to access and inhibit endoplasmic reticulum α-glucosidases, as demonstrated by free oligosaccharide analysis in uninfected and infected SW13 and uninfected Vero cells. Even so, iminosugars may yet have potential as antivirals for CCHFV since the positions and importance of N-linked glycans may differ between the viruses, a hypothesis requiring further evaluation. [ABSTRACT FROM AUTHOR]

Background: Whether prebiotic human milk oligosaccharides (HMO), such as 2'-fucosyllactose (2'-FL) and lacto-N-neotetraose (LNnT), enhance iron absorption in infants is unknown. Moreover, whether maternal HMO profile affects absorption of iron fortificants or the effects of prebiotic galacto-oligosaccharides (GOS) and/or HMO on iron absorption is uncertain. Objectives: The aim of this study was to test whether consumption of 3.0 g GOS or HMO enhances iron absorption from iron-fortified maize porridge in partially breastfed Kenyan infants and whether maternal HMO profile modulates these effects. Methods: In a randomized, prospective crossover study, 55 infants (aged 8-12 mo) were fed test meals fortified with 1 of the following: 1) 5.0 mg iron as 54Fe-labeled ferrous fumarate (FeFum); 2) 5.0 mg iron as 58FeFum and 3.0 g GOS (FeFum+GOS); and 3) 5.0 mg iron as 57FeFum and 2.0 g 2'-FL and 1.0 g LNnT (FeFum+HMO). Fractional iron absorption (FIA) was assessed by erythrocyte incorporation of iron isotopes. HMO profiles were determined by capillary gel electrophoresis with laser-induced florescence detection. Data were analyzed with mixed-effect models, and iron dialyzability was measured in vitro. Results: Of the 55 infants included, 49 were fed as instructed. FIA from the FeFum+GOS group [median (IQR) 22.2% (16.5%-25.9%)] was higher than that from the FeFum group [12.5% (9.5%-20.9%)] (P = 0.005). FIA from the FeFum+HMO group was 13.3% (7.1%-24.4%) and did not differ from the FeFum group (P = 0.923). Maternal HMO profile did not predict FIA or modulate the effects of GOS or HMO on FIA. Iron dialyzability ratios at pH 2 of FeFum+GOS to FeFum and FeFum+HMO to FeFum were 2.1 and 0.9 (P = 0.001 and P = 0.322), respectively. Conclusions: In Kenyan infants consuming FeFum-fortified maize porridge, co-provision of 3.0 g GOS increased FIA by 78%, whereas co-provision of 3.0 g HMO did not affect FIA. Variations in maternal HMO profile, including secretor and Lewis phenotype, did not predict FIA. These data argue against a physiologic role for 2'-FL and LNnT in facilitating iron absorption in infancy. The study was registered at clinicaltrials.gov as NCT04163406 (https://clinicaltrials.gov/ct2/show/NCT04163406). [ABSTRACT FROM AUTHOR]

Raffinose family oligosaccharides (RFOs) have diverse structures and exhibit various biological activities. When using RFOs as prebiotics, their structures need to be identified. If we first knew whether an RFO was classical or non-classical, structural identification would become much easier. Here, we cloned and expressed an α-galactosidase (BF0224) from Bacteroides fragilis which showed strict specificity for hydrolyzing α-Gal-(1 → 6)-Gal linkages in RFOs. BF0224 efficiently distinguished classical from non-classical RFOs by identifying the resulting hydrolyzed oligo- and mono-saccharides with HPAEC-PAD-MS. Using this strategy, we identified a non-classical RFO from Pseudostellaria heterophylla (Miquel) Pax with DP6 (termed PHO-6), as well as a classical RFO from Lycopus lucidus Turcz. with DP7 (termed LTO-7). To characterize these RFO structures, we employed four other commercial or reported α-galactosidases in combination with NMR and methylation analysis. Using this approach, we elucidated the accurate chemical structure of PHO-6 and LTO-7. Our study provides an efficient analytical approach to structurally analyze RFOs. This enzyme-based strategy also can be applied to structural analysis of other glycans. [Display omitted] [ABSTRACT FROM AUTHOR]

Capsular polysaccharides (CPS) of Acinetobacter baumannii is a virulence factor with diverse structures. CPS are produced by the CPS biosynthesis gene cluster in their K locus (KL). However, CPS variations may occur due to insertion of additional genes from external sources, e.g., prophages. Recently, the CPS structure from a clinical isolate, BAL062 which includes KL58 locus, was found to have a pseudaminic acid isomer (8ePse5NAc7NAc) as a result of prophage inserted epaA/epaB genes. Here, we report a CPS structure produced by A. baumannii strain MRSN31468 which also belongs to a KL58 type. The K58 CPS structure was determined by 1D and 2D NMR analysis of the oligosaccharides derived from the CPS by a phage depolymerase, and supported by the sugar composition analysis. The K58 CPS structure has the following tetra saccharide repeating unit. [Display omitted] The K58 CPS differs from the CPS from BAL062 only by replacing 8-epimerized β-8ePse5NAc7NAc with β-Pse5NAc7NAc. Acinetobacter baumannii K58 Capsular polysaccharide. [Display omitted] • Acinetobacter baumannii strain MRSN31468 has K58 capsule polysaccharide with structural similarities to K2 and K98. • The strain MRSN31468 CPS has a similar structure to BAL062, but the Pse is not epimerized at its C-8 position. • A new method for CPS extraction from A. baumannii using 90 % phenol is effective. [ABSTRACT FROM AUTHOR]

A polysaccharide, CZS-0-1, was obtained from the marine green algae Codium fragile using ion-exchange and size-exclusion chromatography. Composition and characteristics analyses showed CZS-0-1 was a sulfated galactoarabinan consisting of arabinose, galactose and a small amount of glucose in a ratio of 9:2:1 with 21% sulfate content and a molecular weight of 810 kDa. Structural properties were determined using desulfation and methylation analyses combined with instrument analysis. The results showed that the backbone of CZS-0-1 was (1 → 3)-β-L-Ara p. Its O -4 and/or O -2 positions showed sulfate modification; additionally, it had 10% of (1 → 3)-β-D-Gal p branches at the O -4 position of the (1 → 3)-β-L-Ara p. The galactose side chains also had sulfate modification at the O -4 or O -6 position. The structure of CZS-0-1 was further confirmed by Top-down analysis of the oligosaccharides after oxidated hydrolysis by mass spectrometry. CZS-0-1 exhibited significant heparin-like anticoagulant activity. It exerted anticoagulant effects by inhibiting FIIa and FXa activities with the presence of heparin cofactors. The anticoagulant activity of CSZ-0-1 was closely related to the molecular weight, and the reduction of molecular weight may lead to a significant decrease in the anticoagulant activity. This study demonstrated that the green algae, Codium fragile can be considered as a useful resource for bioactive polysaccharides. [Display omitted] • A sulfated galactoarabinan CZS-0-1 was extracted from the green tide seaweed Codium fragile. • It had (1 → 3)-β-L-Ara p with sulfate groups at O -4 or O -2 and partially branched at O -4 with β-D-Gal p. • CZS-0-1 exhibited heparin-like anticoagulant activity. [ABSTRACT FROM AUTHOR]

The increasing complexity and ubiquity of food processing and the emergence of fraudulent practices have made effective and reliable methods to authenticate food products of utmost importance. Carbohydrates, with various nutritional functions, are abundant in foods and can serve as potential markers for food authentication. However, the complex and diverse structures and properties of carbohydrates, especially polysaccharides, pose challenges. Nonetheless, significant progress has been made in this area. This paper provides an overview of the utilization of carbohydrates in food authentication since 2000, focusing on strategies involving carbohydrate-based markers, carbohydrate profiles, and carbohydrate-protein interaction-based assays. The analytical techniques, applications, challenges and limitations of these strategies are reviewed and discussed. The findings demonstrate that these strategies offer origin verification, quality assessment, adulteration detection, process control, and food species identification. Notably, oligosaccharide analysis has proven effective in food authentication and remains a promising marker, especially for analyzing intricate matrices. The advances in chromatography separation and mass spectrometry identification of isomers and trace amounts of these compounds have facilitated the discovery of such markers. In conclusion, carbohydrate analysis can play a crucial role in food authentication. Future research and development will make the authentication of carbohydrate-rich foods ever more accurate and efficient. • Carbohydrates can effectively serve as markers in authentication of food rich in carbohydrates. • Advances in chromatography, MS, and derivatization techniques addressed carbohydrate analysis challenges • The use of oligosaccharide-based approaches has demonstrated efficiency in food authentication. [ABSTRACT FROM AUTHOR]

Previous studies showed that crude Houttuynia cordata polysaccharides (CHCP) exerted therapeutic effects on acute lung injury induced by influenza A virus (IAV) in mice. Here, an acidic heteropolysaccharide from H. cordata , named HCPM (Mw , 19.1 kDa), was obtained directly from CHCP using sequential ultrafiltration membrane columns. The backbone of HCPM was consisted of 1, 3, 6-β-Man p , 1, 4-α-Gal p A, 1, 2-α-Rha p , and 1, 2, 4-α-Rha p , with main branches of glucan, arabinan, and galactan substituted at C-3 of 1, 3, 6-β-Man p or C-4 of 1, 2, 4-α-Rha p. The structural information was further validated by oligosaccharide sequencing analysis using UPLC-ESI-MS. Furthermore, HCPM exhibited a potent anti-complementary activity with CH 50 value of 254.1 ± 7.8 μg/mL in vitro and significantly attenuated IAV-induced lung and gut injuries in vivo by inhibiting viral replication, reducing inflammatory responses, and suppressing complement overactivation. These results suggested that HCPM might be a key H. cordata substance for pulmonary infection treatment. [Display omitted] [ABSTRACT FROM AUTHOR]

Purpose: Agaro-oligosaccharides (AGO), hydrolysis products of agarose, is known to have antioxidant and anti-inflammatory properties. Speculating that AGO is effective for preventing aging, we investigated the longevity-supporting effects of AGO and their mechanisms using Caenorhabditis elegans. Methods: Caenorhabditis elegans were fed AGO from young adulthood. The lifespan, locomotory activity, lipofuscin accumulation, and heat stress resistance of the worms were examined. To elucidate mechanisms of AGO-mediated longevity, we conducted comprehensive expression analysis using microarrays. Moreover, we used quantitative real-time PCR (qRT-PCR) to verify the genes showing differential expression levels. Furthermore, we measured the lifespan of loss-of-function mutants to determine the genes related to AGO-mediated longevity. Results: AGO extended the lifespan of C. elegans, reduced lipofuscin accumulation, and maintained vigorous locomotion. The microarray analysis revealed that the endoplasmic reticulum-unfolded protein response (ER-UPR) and insulin/insulin-like growth factor-1-mediated signaling (IIS) pathway were activated in AGO-fed worms. The qRT-PCR analysis showed that AGO treatment suppressed sir-2.1 expression, which is a negative regulator of ER-UPR. In loss-of-function mutant of sir-2.1, AGO-induced longevity and heat stress resistance were decreased or cancelled completely. Furthermore, the pro-longevity effect of AGO was decreased in loss-of-function mutants of abnormal Dauer formation (daf) -2 and daf-16, which are IIS pathway-related genes. Conclusion: AGO delays the C. elegans aging process and extends their lifespan through the activations of ER-UPR and the IIS pathway. [ABSTRACT FROM AUTHOR]

Fabrication and analyses of mucoadhesive patches made from chitosan oligosaccharide for the purpose of oromucosal drug delivery. The mucosal epithelium in the oral cavity, consisting of buccal and sublingual epithelium, has gained significant attention in the last decade as an alternative anatomical site for systemic drug delivery that could potentially minimize the challenges of solid oral dosage and parenteral delivery. In this study, we have fabricated and tested drug-loaded chitosan oligosaccharide-based patches for the oromucosal drug delivery. The chitosan oligosaccharide (with and without alginate) based patches were fabricated using the conventional solvent casting method and were analyzed for their swelling capacity, hydrophilicity, anti-cancer activity, in vitro drug release, and in vivo drug release activity. The in-house developed artificial saliva was used for the swelling study. Alginate-containing patches showed lesser swelling ability compared to the bare chitosan oligosaccharide-based patches. The former was also found to be more hydrophobic compared to the latter one. Both the unloaded patches restricted the growth of epithelial cancer cells indicating their anti-cancer behavior. In vitro drug release indicated a super case II release pattern while in vivo study demonstrated the release of drug from the patch into the plasma indicating the purpose of the fabricated patch. The chitosan oligosaccharide-based mucoadhesive hydrogel patch fabricated in this study can be highly suitable for possible translational purposes. [ABSTRACT FROM AUTHOR]

ALG9-CDG is a CDG-I defect within the group of Congenital Disorders of Glycosylation (CDG). We here describe the clinical symptoms of two new and unrelated ALG9-CDG patients, both carrying the novel homozygous missense variant c.1460 T > C (p.L487P) in the ALG9 gene which led to global developmental delay, psychomotor disability, facial dysmorphisms, brain and heart defects, hearing loss, hypotonia, as well as feeding problems. New clinical symptoms comprised West syndrome with hypsarrhythmia. Quantitative RT-PCR analysis revealed a significantly enhanced ALG9 mRNA transcript level, whereas the protein amount in fibroblasts was significantly reduced. This could be ascribed to a stronger degradation of the mutated ALG9 protein in patient fibroblasts. Lipid-linked oligosaccharide analysis showed an ALG9-CDG characteristic accumulation of Man 6 GlcNAc 2 -PP-dolichol and Man 8 GlcNAc 2 -PP-dolichol in patient cells. The clinical findings of our patients and of all previously published ALG9-CDG patients are brought together to further expand the knowledge about this rare N-glycosylation disorder. SYNOPSIS: Homozygosity for p.L487P in ALG9 causes protein degradation and leads to West syndrome. [ABSTRACT FROM AUTHOR]

The P. heterophylla and its adulterants were identified by HPLC-CAD fingerprint of sucrose and oligosaccharides in P. heterophylla. The improved quantitative analysis of multi-components with a single marker (iQAMS) was further established for simultaneous determinations of sucrose and oligosaccharides in P. heterophylla. The HPLC-CAD fingerprint and similarity coefficients between P. heterophylla and its adulterants showed significant differences. The relative errors (REs) between iQAMS method and external standard method (ESM) were below 3.00%, but significant difference was shown between iQAMS (different marker for whole program with gradient elution) and QAMS (one marker for whole program with gradient elution), indicating that QAMS method should be improved, especially for gradient elution which influence the response of analytes. The accuracy, precision, reproducibility, and stability of this method were validated which exhibited satisfactory results, indicating that iQAMS method could be used for quantitative analysis of sucrose and oligosaccharides in P. heterophylla instead of ESM. The iQAMS combined with HPLC-CAD fingerprint could be used to determine the content of each oligosaccharide, and it can be used for quality control of P. heterophylla. • P. heterophylla and its adulterants were firstly rapid identification by HPLC-CAD fingerprint. • A novel method, HPLC-CAD combined with improved QAMS, was developed for P. heterophylla analysis. • Total contents of sucrose and oligosaccharides were quantified by improved QAMS with PHO DP3–1 and PHO DP4. • The developed QAMS can improve the accuracy of HPLC quantification with gradient elution in the absence of reference standards. [ABSTRACT FROM AUTHOR]

A bioactive polysaccharide (TS2–2A) with a molecular weight of 15 kDa was isolated from Trametes sanguinea Lloyd, a medicinal food homologous fungus, by water extraction-alcohol precipitation and chromatographic separation. NMR analysis of polysaccharide and MS/MS analysis of its oligosaccharide indicated that TS2–2A featured a novel straight chain with a backbone of 1,3-α- d -glucopyranose and 1,4-β- d -glucopyranose at a molar ratio of 1:4. Moreover, TS2–2A, recognized by Toll-like receptor 4 (TLR4), stimulated RAW 264.7 macrophages to release related cytokines and contributed to immune-enhancing effects. Briefly, with remarkable immune-enhancing activity and noncytotoxicity, TS2–2A was proposed to be a potential immune enhancer for supplementing drugs or functional foods. [Display omitted] • Polysaccharide (TS2–2A) was isolated from Trametes sanguinea Lloyd. • TS2–2A mainly comprised glucose with a molecular weight of 15 kDa. • The main structure of TS2–2A was a straight chain with a backbone of →3)-α-D-Glc p -(1 → and →4)-β-D-Glc p -(1 → at a molar ratio of 1:4. • TS2–2A exerted immune-enhancing effects via activation of TLR4. [ABSTRACT FROM AUTHOR]

This study reveals that unexpected degradation of food oligosaccharides can occur during conventional glycomics workflows, including sample preparation and analysis by liquid chromatography-mass spectrometry (LC-MS). With the present investigation, we aim to alert the scientific community of the susceptibility of specific glycosidic linkages to degradation induced by heat and acid. Key standard oligosaccharides representing the major types found in foods (3'-sialyllactose and 6'-sialyl-N-acetyllactosamine for milk, raffinose and stachyose for legumes) were selected as model systems and underwent each of the following treatments independently: (1) labeled with the derivatizing agent 1-aminopyrene-3,6,8-trisulfonic (APTS) (followed by analysis with a capillary electrophoresis system coupled with a fluorescence detector), (2) dried from an acetonitrile-water mixture containing 0.1% trifluoroacetic acid, and (3) injected into an LC-MS system. We demonstrated that both raffinose and stachyose degraded during APTS-labeling by the acid in the labeling reagents. We also discovered that during centrifugal evaporation at 37 °C, all of the four nonderivatized oligosaccharides tested were partially degraded. Additionally, when the LC-MS eluent contained 0.1% formic acid, 3'-sialyllactose, raffinose, and stachyose underwent extensive in-source fragmentation during analysis. Lastly, we identified a simple strategy that can reduce the probability of incorrect oligosaccharide identification resulting from extensive in-source fragmentation. [ABSTRACT FROM AUTHOR]

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Qi-Fu-Yin, a traditional Chinese medicine formula, has been used to treat Alzheimer's disease (AD, a neurodegenerative disorder) in clinical setting. In this study, the chemical components of Qi-Fu-Yin and its prototype components and metabolites in rat plasma and cerebrospinal fluid, after oral administration, were preliminarily characterized via ultrahigh-performance liquid chromatography coupled with quadrupole time-of-flight tandem mass spectrometry (UPLC-Q-TOF-MS). A total of 180 compounds, including saponins, flavonoids, organic acids, sucrose esters, oligosaccharide esters, phthalides, phenylethanoid glycosides, alkaloids, xanthones, terpene lactones, ionones, and iridoid glycoside, were tentatively characterized. For the first time, 51 prototypical components and 26 metabolites, including saponins, phthalides, flavonoids, sucrose esters, organic acids, alkaloids, ionones, terpene lactones, iridoid glycoside, and their derivatives, have been tentatively identified in the plasma. Furthermore, 10 prototypical components (including butylidenephthalide, butylphthalide, 20(S)-ginsenoside Rh1, 20(R)-ginsenoside Rh1, and zingibroside R1) and 6 metabolites were preliminarily characterized in cerebrospinal fluid. These results were beneficial to the discovery of the active components of Qi-Fu-Yin anti-AD. [ABSTRACT FROM AUTHOR]

Mammalian hyaluronidases are endo- N -acetyl-D-hexosaminidases involved in the catabolism of hyaluronic acid (HA) but their role in the catabolism of chondroitin sulfate (CS) is also examined. HA and CS are glycosaminoglycans implicated in several physiological and pathological processes, and understanding their metabolism is of significant importance. Data have been previously reported on the degradation of CS under the action of hyaluronidase, yet a detailed structural investigation of CS depolymerization products remains necessary to improve our knowledge of the CS depolymerizing activity of hyaluronidase. For that purpose, the fine structural characterization of CS oligosaccharides formed upon the enzymatic depolymerization of various CS subtypes by hyaluronidase has been carried out by high-resolution Orbitrap mass spectrometry (MS) and extreme UV (XUV) photodissociation tandem MS. The exact mass measurements show the formation of wide size range of even oligosaccharides upon digestion of CS-A and CS-C comprising hexa- and octa-saccharides among the main digestion products, as well as formation of small quantities of odd-numbered oligosaccharides, while no hyaluronidase activity was detected on CS-B. In addition, slight differences have been observed in the distribution of oligosaccharides in the digestion mixture of CS-A and CS-C, the contribution of longer oligosaccharides being significantly higher for CS-C. The sequence of CS oligosaccharide products determined XUV photodissociation experiments verifies the selective β(1 → 4) glycosidic bond cleavage catalyzed by mammal hyaluronidase. The ability of the mammal hyaluronidase to produce hexa- and higher oligosaccharides supports its role in the catabolism of CS anchored to membrane proteoglycans and in extra-cellular matrix. [ABSTRACT FROM AUTHOR]

• Caprine and bovine dairy products had distinguished oligosaccharide profiles. • Oligosaccharide profiles in adulterated caprine dairy products were analyzed. • LNTri is a potential marker distinguishing caprine and bovine dairy products. Adulteration of caprine dairy products raises concerns among consumers. This study aimed to identify the differences in oligosaccharide profiles of caprine dairy products, including raw milk, colostrum powder, and lactose powder, and their corresponding bovine dairy products, and provide new insights for detecting adulteration of bovine dairy products in caprine dairy products. Twenty-seven oligosaccharides were detected in caprine and bovine dairy products. The principal component analysis plot of the oligosaccharide profiles clearly differentiated among the six types of dairy products. Specific oligosaccharides that were most distinctive for caprine and bovine dairy products were identified. Lacto- N -triose (LNTri) could be used as a potential biomarker for distinguishing caprine milk from bovine milk, caprine colostrum powder from bovine colostrum powder, and caprine lactose powder from bovine lactose powder. The results demonstrated that oligosaccharides could be used as biomarkers for detecting bovine dairy products in caprine dairy products, especially caprine lactose powder. [ABSTRACT FROM AUTHOR]

Background: Human milk oligosaccharides (HMOs) are complex glycans that are highly abundant in human milk. While over 150 HMOs have been identified, it is unknown how individual HMOs change in concentration over 24 months of lactation.Objectives: To understand how HMO concentrations change over 24 months of lactation.Methods: Breast milk samples were collected from participants in a longitudinal cohort study of Hispanic mother-infant pairs at 1, 6, 12, 18, and 24 months postpartum. Concentrations of 19 of the most abundant HMOs were measured using HPLC. Because the parent study is ongoing and not all participants have finished all time points yet, the sample sizes ranged per time point (n = 207 at 1 month; n = 109 at 6 months; n = 83 at 12 months; n = 59 at 18 months; and n = 28 at 24 months). Approximately 88% of participants were classified as HMO secretors-a genetic factor that affects concentrations of HMOs such as 2'fucosyllactose (2'FL) and lacto-N-fucopentaose I-while the remaining 12% were classified as nonsecretors. Mixed models were used to examine changes in HMO concentrations and relative abundances over the course of lactation.Results: The majority of HMOs significantly decreased in concentration over the course of lactation. The exceptions were 2'FL, sialyl-lacto-N-tetraose b, and disialyl-lacto-N-tetraose, which did not change with time, and 3-fucosyllactose (3FL) and 3'-sialyllactose (3'SL), which significantly increased. The concentration of 3FL increased 10-fold, from 195 (IQR 138-415) μg/mL at 1 month to 1930 (1100-2630) μg/mL at 24 months, while 3'SL increased 2-fold, from 277 (198-377) μg/mL to 568 (448-708) μg/mL over the same time period.Conclusions: These results indicate that HMOs do not decrease in concentration uniformly across lactation. In particular, 3FL and 3'SL increased over the course of lactation in this cohort. Future studies are required to fully understand the functions of these HMOs. [ABSTRACT FROM AUTHOR]

BACKGROUND The present study aimed to explore the chemical characteristics of mountainous forest cultivated ginseng (MFCG) and garden ginseng (GG) with respect to their ginsenosides and oligosaccharides. METHODS: A high‐performance liquid chromatography with diode‐array detection‐evaporative light‐scattering detection technique was adopted to investigate the ginseosides and oligosaccharides of GG and MFCG. RESULTS: The features of ginsenosides showed Rg1/Re in different parts of GG and MFCG: main root > lateral root > fibrous root, as well as Rg1/Re in the main root: MFCG > GG, indicating that the Rg1/Re is related to age of the ginseng. In most cases, Rg1/Re < 1 in entire GG and Rg1/Re > 1 in entire MFCG. In addition, the ratio of protopanaxadiol/protopanaxatriol in main root of GG is approximately 1 and, in the main roots of MFCG, the ratio is approximately 2 and, furthermore, Ro/Rb1 of MFCG is lower than that of GG. Analysis of oligosaccharides showed that GG mainly contained sucrose and MFCG mainly contained sucrose and maltose, and the ratio of sucrose to maltose was at least more than 4:1 in GG and less than 4:1 in MFCG in most cases, indicating the characteristics of oligosaccharides of MFCG are primarily affected by its growing environment. The results also showed that ginsenoside Re is most probably the biosynthetic precursor of ginsenoside Rg1 (i.e. Re was synthesized first and then transformed to Rg1 in vivo). CONCLUSION: The characteristics of Rg1/Re and higher maltose can be regarded as one of the characteristics of high quality MFCG, and these characteristics are related to a higher age and the cultivation environment of ginseng. The formation mechanism of these characteristics for GG and MFGG is also discussed. As far as we know, the present study is the first to determine the difference of Rg1/Re and oligosaccharides between MFCG and GG and this provides a reference for the quality control criterion of GG and MFCG. © 2020 Society of Chemical Industry [ABSTRACT FROM AUTHOR]

MOGS mutations cause congenital disorders of glycosylation type IIb (CDG-IIb or GCS1-CDG). The specific manifestations caused by the mutations in this gene remain unknown. We aimed to describe the clinical features of CDG- IIb and the effectiveness of urinary oligosaccharide analysis in the diagnosis of CDG- IIb. Patient 1 was analyzed with whole-exome sequencing (WES) to identify the causative gene of intractable epilepsy and severe developmental delay. After detecting MOGS mutation in patient 1, we analyzed patients 2 and 3 who were siblings and had clinical features similar to those in patient 1. Urinary oligosaccharide analysis was performed to confirm CDG- IIb diagnosis in patient 1. The clinical features of these patients were analyzed and compared with those in eight published cases. Our three patients presented with early infantile epileptic encephalopathy, generalized hypotonia, hepatic dysfunction and dysmorphic features. In two cases, compound heterozygous mutations in MOGS were identified by WES. Isolation and characterization of the urinary oligosaccharide was performed in one of these cases to confirm the diagnosis of CDG-IIb. Although the isoelectric focusing of transferrin (IEF-T) of serum in this patient was normal, urinary excretion of Hex 4 corresponding to Glc 3 Man was observed by mass spectrometry. This report provides clinical manifestations of CDG-IIb with MOGS mutation. CDG-IIb shows a normal IEF profile of serum transferrin and cannot be detected by structural analysis of the patient's glycoproteins. Characterization of urinary oligosaccharides should be considered to detect this disorder. [ABSTRACT FROM AUTHOR]

Lactic acid bacteria (LAB) are important in food fermentation and may enhance overall host health. Previous studies to explore LAB metabolism mainly focused on the genera Lacticaseibacillus and Lactococcus. Pediococcus pentosaceus, historically recognized as an important food fermentation bacterial strain, can produce bacteriocins and occasionally demonstrated probiotic functionalities. This study thoroughly surveyed the growth kinetic of three P. pentosaceus isolates in various culture formulations, especially in fructooligosaccharide (FOS), xylooligosaccharide (XOS), or konjac mannooligosaccharide (KMOS) conditions. Results showed that P. pentosaceus effectively metabolized KMOS, the culture of which led to 23.6-fold population increase. However, FOS and XOS were less metabolized by P. pentosaceus. On functional oligosaccharide cultures, P. pentosaceus could result in higher population proliferation, more acidified fermentation environment, and higher glycoside hydrolysis activities in the culture. RNA-Seq analysis classified 1572 out of 1708 putative genes as mRNA-coding genes. The dataset also revealed that the three functional oligosaccharides led to extensive global functional gene regulations. Phosphate conservation and utilization efficiency enhancement may serve as a leading transcriptional regulation direction in functional oligosaccharide metabolisms. In summary, these discovered metabolic characteristics could be employed to support future studies. Key points: • Konjac mannooligosaccharides effectively promoted P. pentosaceus proliferation. • Functional genes were highly regulated in functional oligosaccharide utilization. • Phosphate conservation was an important transcriptional regulation direction. [ABSTRACT FROM AUTHOR]

This study reports on the development of a novel instrument for capillary electrophoresis (CE) coupled with laser induced fluorescence (LIF) detection that is inspired by the Lego-toy concept. The Lego CE-LIF design is an evolution of purpose-made CE instrumentation, allowing the users to construct their own analytical device with a high degree of standardization (i.e. a "standard" setup) without requirement of mechanical and electronic workshop facilities. To allow instrument reproduction outside the original fabrication laboratory, which is not trivial for in-house-built CE systems, the new design is based on unprecedent 'plugging' hyphenation of various off-the-shelf parts available for microfluidics, optics and electrophoresis. To render the operation with Lego CE-LIF optimal, we developed a new background electrolyte (BGE), using for the first time extremely high concentrations of zwitterionic and large weakly charged species for much improvement of detection sensitivity. The Lego CE-LIF was demonstrated for separation and detection of oligosaccharides labelled with 8-aminopyrene-1,3,6-trisulfonic acid (APTS). The new gel-free BGE for oligosaccharide analysis also allowed simplification of the conventional CE-LIF protocol used with commercial instruments while keeping satisfactory separation performances. Furthermore, the new BGE is fully compatible with a non-thermostatted Lego CE instrument thanks to low current and therefore low heat generation under application of a high voltage. Lego-toy-inspired instrumentation for capillary electrophoresis with laser induced fluorescence detection. Image 1 • A novel instrumental design inspired by the Lego-toy concept was developed. • A Lego-CE instrument was constructed from off-the-shelf microfluidic and electrophoretic components. • A Lego-LIF detector was developed from a USB-powered laser source, ready-to-use microfluidic and optic parts. • A new background electrolyte was developed for improvement of CE-LIF performance. • Different modes of pressure-assisted electrophoresis was demonstrated with the Lego-CE-LIF system. [ABSTRACT FROM AUTHOR]

Polysaccharides are the most abundant biomolecules in nature, but are the least understood in terms of their chemical structures and biological functions. Polysaccharides cannot be simply sequenced because they are often highly branched and lack a uniform structure. Furthermore, large polymeric structures cannot be directly analyzed by mass spectrometry techniques, a problem that has been solved for polynucleotides and proteins. While restriction enzymes have advanced genomic analysis, and trypsin has advanced proteomic analysis, there has been no equivalent enzyme for universal polysaccharide digestion. We describe the development and application of a chemical method for producing oligosaccharides from polysaccharides. The released oligosaccharides are characterized by advanced liquid chromatography–mass spectrometry (LC–MS) methods with high sensitivity, accuracy and throughput. The technique is first used to identify polysaccharides by oligosaccharide fingerprinting. Next, the polysaccharide compositions of food and feces are determined, further illustrating the utility of technique in food and clinical studies. While mass spectrometry-based proteomics largely relies on digesting proteins into peptides, there is no equivalent strategy for polysaccharide analysis. Here, the authors develop a chemical approach to break down poly- into oligosaccharides and present a workflow to identify polysaccharides by oligosaccharide fingerprinting. [ABSTRACT FROM AUTHOR]

Complete heparin digestion with heparin lyase I and II results in a mixture of hexasaccharides and tetrasaccharides with 3‐O‐sulfo group‐containing glucosamine residues at their reducing ends. Because these tetrasaccharides are derived from antithrombin III‐binding sites of heparin, we examined whether this method could be applied to estimate the anticoagulant activity of heparin. Therefore, this paper presents a new low molecular weight heparin sample preparation method–chemical depolymerization. Qualitative analysis of the studied compounds and a comparison of their composition are an important contribution to the structural analysis of low molecular weight heparins, which has not been fully conducted so far. Qualitative on‐line liquid chromatography–mass spectrometric analysis of these resistant oligosaccharides is also described in this paper. [ABSTRACT FROM AUTHOR]

Introduction: Introduction: human milk oligosaccharides (HMOs) are an important component of human milk supporting the development of a balanced intestinal microbiota and immune protection in breastfed infants. Randomized controlled trials (RCTs) have demonstrated that infant formulas supplemented with the HMOs 2'-fucosyllactose (2'FL) and lacto-N-neotetraose (LNnT) are safe, well-tolerated, and support normal growth. This Real-World Evidence (RWE) study aimed to evaluate growth and tolerance in infants consuming a formula supplemented with 1 g/L of 2'FL and 0.5 g/L of LNnT, and included a mixed-feeding group never studied before in RCTs. Participants and methods: this open-label, prospective study was conducted at six centers in Spain, and included healthy, exclusively breastfed infants (BF group), an exclusively formula-fed group (FF) who received a milk-based formula with 2' FL and LNnT, and a group mixed fed with both formula and human milk (MF), for 8 weeks. Co-primary outcomes were growth (anthropometry) and gastrointestinal tolerance (Infant Gastrointestinal Symptom Questionnaire, IGSQ). Secondary outcomes included formula satisfaction and adverse events (AEs). Results: 159 infants completed the study (66 FF, 48 MF, and 45 BF). Mean z-scores for growth were similar between all groups and within ± 0.5 of WHO medians at week 8. Composite IGSQ scores demonstrated low GI distress in all groups, with no significant group differences at baseline, week 4, or week 8. Incidence of AEs was low overall, and comparable across groups. Conclusions: in this RWE study examining a HMO-supplemented infant formula, growth and tolerance outcomes were similar to RCT findings, supporting the effectiveness of this early feeding option. [ABSTRACT FROM AUTHOR]

[Objective] In order to systematically evaluate the effect of functional oligosaccharides on immune organ index in broilers. [Method] CNKI database was searched by computer, and the related literature was screened by the inclusion standard, screened relevant literatures through inclusion criteria, and Meta analyzed 29 studies of 13 literatures obtained. [Result] Compared with the control group, the thymus index of the oligosaccharide group was increased [WMD =0. 243,95 % CI = (0. 173 ,0. 314) ,P =0. 000], bursa index [WMD =0.097 ,95 % CI = (0. 072,0.122) ,P = 0. 000 ] ,spleen index [WMD = 0. 094,95 % CI = (0. 067 ,0. 121) , P = 0. 000] were significantly higher than those of the control group. The results of subgroup analysis based on oligosaccharide additives showed that the effects of different types of functional oligosaccharides on immune organ index of broilers were not consistent and the effects of fruit oligosaccharides on thymus index and pectin oligosaccharides and zinc coordient on the index of spleen and fasciae were the best. The results of sensitivity analysis showed that adding functional oligosaccharides to diet significantly increased the immune organ index of broilers. There was no significant publication bias in the studies on thymus index and bursa index, but there was some publication bias in the studies on spleen index. [Conclusion] Based on the existed evidence, dietary supplementation of functional oligosaccharide could increase the immune organ index of 42-day-old broilers. [ABSTRACT FROM AUTHOR]

Background Oligosaccharides are the third most abundant component in human milk. They are a potential protective agent against neonatal sepsis. Objectives We aimed to explore the association between human milk oligosaccharides (HMOs) and late-onset sepsis in very-low-birth-weight infants, and to describe the composition and characteristics of HMOs in Peruvian mothers of these infants. Methods This is a secondary data analysis of a randomized clinical trial. We conducted a retrospective cohort study of mothers and their very-low-birth-weight (<1500 g) infants with ≥1 milk sample and follow-up data for >30 d. HMOs were measured by high performance liquid chromatography (HPLC). We used factor analysis and the Mantel–Cox test to explore the association between HMOs and late-onset neonatal sepsis. Results We included 153 mother–infant pairs and 208 milk samples. Overall, the frequency of the secretor phenotype was 93%. Secretors and nonsecretors were defined by the presence and near-absence of α1-2-fucosylated HMOs, respectively. The most abundant oligosaccharides were 2'-fucosyllactose, lacto-N-fucopentaose (LNFP) I, and difucosyllacto-N-tetraose in secretors and lacto-N-tetraose and LNFP II in nonsecretors. Secretors had higher amounts of total oligosaccharides than nonsecretors (11.45 g/L; IQR: 0.773 g/L compared with 8.04 g/L; IQR: 0.449 g/L). Mature milk samples were more diverse in terms of HMOs than colostrum (Simpson's Reciprocal Diversity Index). We found an association of factor 3 in colostrum with a reduced risk of late-onset sepsis (HR: 0.63; 95% CI: 0.41, 0.97). Fucosyl-disialyllacto-N-hexose (FDSLNH) was the only oligosaccharide correlated to factor 3. Conclusions These findings suggest that concentrations of different HMOs vary from one individual to another according to their lactation period and secretor status. We also found that FDSLNH might protect infants with very low birth weight from late-onset neonatal sepsis. Confirming this association could prove 1 more mechanism by which human milk protects infants against infections and open the door to clinical applications of HMOs. This trial was registered at clinicaltrials.gov as NCT01525316. [ABSTRACT FROM AUTHOR]

This paper addresses two fundamental issues in the peak deconvolution method of cellulose XRD data analysis: there is no standard model for amorphous cellulose and common peak functions such as Gauss, Lorentz and Voigt functions do not fit the amorphous profile well. It first examines the effects of ball milling on three types of cellulose and results show that ball milling transforms all samples into a highly amorphous phase exhibiting nearly identical powder X-ray diffraction (XRD) profiles. It is hypothesized that short range order within a glucose unit and between adjacent units survives ball milling and generates the characteristic amorphous XRD profiles. This agrees well with cellulose I d-spacing measurements and oligosaccharide XRD analysis. The amorphous XRD profile is modeled using a Fourier series equation where the coefficients are determined using the nonlinear least squares method. A new peak deconvolution method then is proposed to analyze cellulose XRD data with the amorphous Fourier model function in conjunction with standard Voigt functions representing the crystalline peaks. The impact of background subtraction method has also been assessed. Analysis of several cellulose samples was then performed and compared to the conventional peak deconvolution methods with common peak fitting functions and background subtraction approach. Results suggest that prior peak deconvolution methods overestimate cellulose crystallinity. [ABSTRACT FROM AUTHOR]

Oligosaccharides are one of the functional ingredients to be used in food technology. In this study, by using an active glucansucrase GTFA-ΔN E81 in the acceptor reaction of mannose, mannose-containing oligosaccharides were produced and their functionality was tested. The formation of the oligosaccharides were visualised by TLC analysis and mannose-containing oligosaccharides up to DP 7 were determined by LC-MS analysis. The presence of the (1,6)Glc and (1,3)Glc units within the oligosaccharides were determined by NMR analysis. The in vitro immune-modulatory functions of the mannose-containing oligosaccharides were determined but no induction in IL-4, IL-10, IL-12 and TNF-α cytokine levels were detected. Importantly this oligosaccharide mixture showed prebiotic effect by triggering the growth of tested probiotics and not affecting the growth of pathogen strains. Our findings reveals the potential of the role of glucansucrases for the production of functional oligosaccharides. [ABSTRACT FROM AUTHOR]

Oxidized starch (oxStarch) is a major derivative of starch. In present study, 4-acetamide-TEMPO system was firstly applied to prepare specifically oxidized starch, homogeneous 1,4-linked α-D-glucuronan. The impact of oxidant amount, 4-acetamide-TEMPO amount and reaction temperature on the properties of products were investigated. The product structures were characterized with 1H NMR for degree of oxidation (DO), SEC-MALS for MW, MS for analysis of oxidized oligosaccharides, and in-source fragmented MS for analysis of oxidized polysaccharides. The results showed that the DO of oxStarches increased, but their MWs decreased with the raise of oxidant amount; higher 4-acetamide-TEMPO amount provided higher efficiency of specific oxidation of starch, in which the product has higher DO and MW; no significant difference have been observed between the oxStarches oxidized at 4 and 25 °C, but significant degradation was observed at 50 °C. In each product, having a MW distribution, the portion with smaller size has higher DO. While some unspecific oxidation was still observed observed on the second hydroxyl groups of sugar ring in the following order of priority, position 3 > 2 > 1 > 0. [ABSTRACT FROM AUTHOR]

Intestinal microbiota plays an important role in human health. The aim of this paper is to determine the impact of the phenolics and carbohydrate in buckwheat honey on human intestinal microbiota. We investigated the phenolics and carbohydrate compositions of eight buckwheat honey samples using high-performance liquid chromatography and ion chromatography. The human intestinal microbes were cultured in a medium supplemented with eight buckwheat honey samples or the same concentration of fructooligosaccharides. The bacterial 16S rDNA V4 region sequence of DNA extraction was determined by the Illumina MiSeq platform. 12 phenolics and 4 oligosaccharides were identified in almost all buckwheat honey samples, namely, protocatechuic acid, 4-hydroxy benzoic acid, vanillin, gallic acid, p-coumaric acid, benzoic acid, isoferulic acid, methyl syringate, trans,trans-abscisic acid, cis,trans-abscisic acid, ferulic acid, 4-hydroxybenzaldehyde, kestose, isomaltose, isomaltotriose, and panose. Most notably, this is the first study to reveal the presence of 4-hydroxybenzaldehyde in buckwheat honey. 4-Hydroxybenzaldehyde seems to be a land marker of buckwheat honey. Our results indicate that buckwheat honey can provide health benefits to the human gut by selectively supporting the growth of indigenous Bifidobacteria and restraining the pathogenic bacterium in the gut tract. We infer that buckwheat honey may be a type of natural intestinal-health products. [ABSTRACT FROM AUTHOR]

Yeast α-mannan (YM) is a densely branched N-linked glycan that decorates the surface of yeast cell walls. Owing to the high degree of branching, cleavage of the backbone of YM appears to rely on the coupled action of side-chain-cleaving enzymes. Upon examining the genome sequences of bovine-adapted Bacteroides thetaiotaomicron strains, isolated for their ability to degrade YM, we have identified a tandem pair of genes inserted into an orphan pathway predicted to be involved in YM metabolism. Here, we investigated the activity of one of these enzymes, a predicted endo-mannanase from glycoside hydrolase (GH) family 76 (BtGH76-MD40). Purified recombinant BtGH76-MD40 displayed activity on structurally distinct YMs from Saccharomyces cerevisiae and Schizosaccharomyces pombe. Linkage analysis of released oligosaccharide products from S. cerevisiae and S. pombe mannan determined BtGH76-MD40 targets a specific linkage that is conserved in structurally diverse YM substrates. In addition, using two differential derivatization methods, we have shown that there is an absolute requirement for undecorated d -mannopyranose in the −1 subsite. Determination of the BtGH76-MD40 X-ray crystal structure and structural superimposition and molecular docking of a branched alpha-mannopentatose substrate supported these findings. In contrast, BtGH76-MD40 can accommodate extended side chains in the +1 and −2 subsites, highlighting that a single alpha-1,6-mannosyl residue is a prerequisite for activity, and cleavage occurs at the reducing end of the undecorated monosaccharide. Collectively these results demonstrate how acquisition of new enzymes within extant pathways contributes to the functional abilities of saccharolytic bacteria persisting in complex digestive ecosystems. Image 1 • Yeast mannan (YM) is a highly branched polysaccharide with species-dependent structural diversity. • An enzyme has been found within the bovine rumen that cleaves the backbone of YM. • Cleavage requires the presence of an unbranched 6-linked mannose. • Glycosidic linkage analysis and crystallography elucidate the nature of interactions. [ABSTRACT FROM AUTHOR]

In this work, new multi-cationic aminopyrene-based labeling tags were designed and synthesized for oligosaccharide analysis by capillary electrophoresis-mass spectrometry (CE-MS). The starting compound, 8-aminopyrene-1,3,6-trisulfonic acid trisodium salt, was modified in order to form a sulfonamide derivative having three tertiary amines in the label structure. The sulfonamide derivative was further methylated to generate three permanently charged quaternary ammonium moieties on the label. The synthesized labels were characterized by NMR, IR, UV/Vis, fluorescence spectroscopy and mass spectrometry. Furthermore, the labels were applied for maltooligosaccharide standards as well as N -linked glycans labeling via reductive amination and followed by CE-MS analysis. The CE-MS analysis of maltooligosaccharides labeled by these newly synthesized labels provided the sub-micromolar limit of detection based on the extracted ion electropherogram signals. Image 1 • New multi-cationic aminopyrene-based labeling tags were designed and synthesized. • The synthesized labels were characterized by NMR, IR, UV/Vis, fluorescence spectroscopy, and mass spectrometry. • The labels were successfully applied for maltooligosaccharides and N -linked glycans labeling followed by CE-MS analysis. [ABSTRACT FROM AUTHOR]