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Sample records for fructo oligosaccharides production

  1. Fructo-oligosaccharide production from inulin through partial citric or phosphoric acid hydrolyses.

    PubMed

    Fontana, José Domingos; Grzybowski, Adelia; Tiboni, Marcela; Passos, Maurício

    2011-11-01

    Purified inulin from Dahlia tubers was partially hydrolyzed to form fructo-oligosaccharides by using citric or phosphoric acids (pH, 2.0-2.5) as mild acid catalysts. The ideal kinetic conditions to ensure a high yield of fructo-oligosaccharides relative to free fructose were a temperature range of 85°C-95°C, a hydrolysis time of 15-25 minutes, and a catalyst pH of 2.5. At the higher temperature and the longest hydrolysis time, an inversion of the product ratio occurred. Under these conditions, co-generation of hydroxymethylfurfural occurred, and it was eliminated by activated charcoal. Unlike in classic hydrolysis with hydrochloric or sulfuric acid, deionization of the actual hydrolysates was not necessary because the catalyst neutralization with common bases results in the formation of co-nutrients with alternative uses as foods or fermentation substrates. These whole hydrolysates can be advantageously added as nutraceuticals to carbonated beverages and acidic foods, such as soft drinks and yogurts.

  2. High-efficient production of fructo-oligosaccharides from inulin by a two-stage bioprocess using an engineered Yarrowia lipolytica strain.

    PubMed

    Han, Yao-Zu; Zhou, Chao-Chao; Xu, Yan-Yan; Yao, Jing-Xin; Chi, Zhe; Chi, Zhen-Ming; Liu, Guang-Lei

    2017-10-01

    A convenient and efficient two-stage bioprocess was established for fructo-oligosaccharides (FOSs) production, during which the endo-inulinase was first produced and subsequently the inulin supplemented was directly hydrolyzed by the produced endo-inulinase, in the meantime the generated non-prebiotic saccharides was assimilated by the yeast cells. This process was implemented by an engineered Yarrowia lipolytica strain Enop56, in which an optimized endo-inulinase gene from Aspergillus niger was overexpressed. When the strain Enop56 was fermented with an inulin concentration of 600g/L in a 10-L bioreactor, the inulinase activity, FOSs titer, yield and productivity reached to 551.6U/mL, 546.6g/L, 0.91gFOS/gInulin, and 15.18g/L/h, respectively. Besides, the hydrolysis products were mainly FOSs with polymerization degrees of 3-5 and the total amount of non-prebiotic mono- and disaccharides was only 4.97% in the final fermentation broth. This study demonstrated that the two-stage bioprocess using the strain Enop56 was a promising strategy to produce FOSs on an industrial scale. Copyright © 2017 Elsevier Ltd. All rights reserved.

  3. Gaseous response to ingestion of a poorly absorbed fructo-oligosaccharide sweetener.

    PubMed

    Stone-Dorshow, T; Levitt, M D

    1987-07-01

    Fructo-oligosaccharides are naturally occurring sweet substances that are poorly absorbed and have the potential to be clinically useful nonnutritive sweeteners. Because most nonabsorbed carbohydrates are fermented yielding gas, we assessed flatulent symptoms and H2 excretion during ingestion of fructo-oligosaccharide (5 g tid) for 12 d. Ten subjects had significantly greater flatulence while taking the oligosaccharide than did five subjects taking sucrose (5 g tid). Breath H2 after 10 g fructo-oligosaccharide was similar to that of 10 g lactulose, suggesting near total malabsorption of the fructo-oligosaccharide. Although previous studies found a marked diminution in breath H2 after prolonged exposure to lactulose, breath H2 response increased by 50% after a 12-d period on the oligosaccharide and gaseous symptoms did not improve. We conclude that adaptation of colonic bacteria to carbohydrate malabsorption is variable and may depend upon quantity or nature of the carbohydrate.

  4. Identification of fructo- and malto-oligosaccharides in cured tobacco leaves (Nicotiana tabacum).

    PubMed

    Nagai, Atsushi; Yamamoto, Takeshi; Wariishi, Hiroyuki

    2012-07-04

    Fructo-oligosaccharides (FOSs) and malto-oligosaccharides (MOSs) in cured tobacco leaves ( Nicotiana tabacum ) were detected and quantified using high-performance liquid chromatography (HPLC) coupled with electrospray ionization (ESI) tandem mass spectrometry (MS/MS). These oligosaccharides were present in several tobacco materials including flue-cured tobacco, sun/air-cured tobacco, and cut filler of commercially available tobacco products, but were not detected in air-cured tobacco. The changes in these oligosaccharides during storage were also investigated. The results revealed that MOSs simply decreased under a warm atmosphere and FOSs increased. In addition, the formation of FOSs in cured tobacco leaves occurred in the presence of sucrose. With regard to FOSs, it has been reported that green tobacco leaves do not contain FOSs such as kestose, nystose, and fructosyl-nystose. The results of a model test suggested that the changes in FOS amount were caused by enzymatic reactions.

  5. Comparison of Yacon (Smallanthus sonchifolius) Tuber with Commercialized Fructo-oligosaccharides (FOS) in Terms of Physiology, Fermentation Products and Intestinal Microbial Communities in Rats

    PubMed Central

    UTAMI, Ni Wayan Arya; SONE, Teruo; TANAKA, Michiko; NAKATSU, Cindy H; SAITO, Akihiko; ASANO, Kozo

    2013-01-01

    The yacon (Smallanthus sonchifolius) tuber was examined with regard to its prebiotic effects compared with commercialized fructo-oligosaccharides (FOS). A feed containing 10% yacon tuber, which is equivalent to 5% commercialized FOS in terms of the amount of fructo-oligosaccharides (GF2, GF3 and GF4), was administrated to rats for 28 days. The yacon diet changed the intestinal microbial communities beginning in the first week, resulting in a twofold greater concentration of cecal short-chain fatty acids (SCFAs). The SCFA composition differed, but the cecal pH in rats fed yacon tuber was equal to that in rats fed FOS. Serum triglycerides were lower in rats fed yacon compared with rats fed FOS and the control diet. Cecal size was greater with the yacon tuber diet compared with the control diet. The abundant fermentation in the intestines created a selective environment for the intestinal microbiota, which included Lactobacillus acidophilus, Bifidobacterium pseudolongum, Bifidobacterium animalis and Barnesiella spp. according to identification with culture-independent analysis, 16S rRNA gene PCR-DGGE combined with cloning and sequencing. Barnesiella spp. and B. pseudolongum were only found in the rats fed the yacon diet, while L. acidophilus and B. animalis were found in abundance in rats fed both the yacon and FOS diets. The genus Barnesiella has not previously been reported to be associated with yacon or FOS fermentation. We concluded that the physiological and microbiological effects of the yacon tuber were different from those of FOS. Differences in cecal size, blood triglycerides and microbial community profiles including their metabolites (SCFAs) between the yacon tuber and FOS were shown to be more greatly affected by the yacon tuber rather than FOS. PMID:24936376

  6. Comparison of Yacon (Smallanthus sonchifolius) Tuber with Commercialized Fructo-oligosaccharides (FOS) in Terms of Physiology, Fermentation Products and Intestinal Microbial Communities in Rats.

    PubMed

    Utami, Ni Wayan Arya; Sone, Teruo; Tanaka, Michiko; Nakatsu, Cindy H; Saito, Akihiko; Asano, Kozo

    2013-01-01

    The yacon (Smallanthus sonchifolius) tuber was examined with regard to its prebiotic effects compared with commercialized fructo-oligosaccharides (FOS). A feed containing 10% yacon tuber, which is equivalent to 5% commercialized FOS in terms of the amount of fructo-oligosaccharides (GF2, GF3 and GF4), was administrated to rats for 28 days. The yacon diet changed the intestinal microbial communities beginning in the first week, resulting in a twofold greater concentration of cecal short-chain fatty acids (SCFAs). The SCFA composition differed, but the cecal pH in rats fed yacon tuber was equal to that in rats fed FOS. Serum triglycerides were lower in rats fed yacon compared with rats fed FOS and the control diet. Cecal size was greater with the yacon tuber diet compared with the control diet. The abundant fermentation in the intestines created a selective environment for the intestinal microbiota, which included Lactobacillus acidophilus, Bifidobacterium pseudolongum, Bifidobacterium animalis and Barnesiella spp. according to identification with culture-independent analysis, 16S rRNA gene PCR-DGGE combined with cloning and sequencing. Barnesiella spp. and B. pseudolongum were only found in the rats fed the yacon diet, while L. acidophilus and B. animalis were found in abundance in rats fed both the yacon and FOS diets. The genus Barnesiella has not previously been reported to be associated with yacon or FOS fermentation. We concluded that the physiological and microbiological effects of the yacon tuber were different from those of FOS. Differences in cecal size, blood triglycerides and microbial community profiles including their metabolites (SCFAs) between the yacon tuber and FOS were shown to be more greatly affected by the yacon tuber rather than FOS.

  7. Production of levansucrase from Bacillus subtilis NRC 33a and enzymic synthesis of levan and Fructo-Oligosaccharides.

    PubMed

    Abdel-Fattah, Ahmed F; Mahmoud, Doaa A R; Esawy, Mona A T

    2005-12-01

    Bacillus subtilis NRC 33a was able to produce both inducible and constitutive extracellular levansucrase, respectively, using sucrose and glucose as carbon source. The optimal production of the levansucrase was at 30 degrees C. The effect of different nitrogen sources showed that baker's yeast with 2% concentration gave the highest levansucrase activity. Addition of 0.15 g/L MgSO(4) was the most favorable for levansucrase production. The enzymic synthesis of levan was studied using 60% acetone fraction. The results indicated that high enzyme concentrations produced increasing amounts of levan, and hence conversion of fructose to levan reached 84% using 1,000 microg/ml enzyme protein. Sucrose concentration was the most effective factor controlling the molecular weight of the synthesized levan. The conversion of fructose to levan was maximal at 30 degrees C. The time of reaction clearly affected the conversion of fructose to levan, which reached its maximum productivity at 18 hours (92%). Identification of levan indicated that fructose was the building unit of levan.

  8. Evaluation of commercial resins for fructo-oligosaccharide separation.

    PubMed

    Nobre, C; Suvarov, P; De Weireld, G

    2014-01-25

    Fructo-oligosaccharides (FOS) produced by fermentative processes are obtained in mixtures containing significant amounts of salts and other non-prebiotic sugars. A demineralisation process using a mixture of a cationic and an anionic resin was proposed. The separation of FOS from a mixture of fructose, glucose and sucrose was evaluated. Experiments were conducted with several commercial cationic exchange resins in calcium, sodium and potassium forms packed in preparative columns (7cm×2.2cm length×diameter). Resins in potassium form obtained the higher retention factor values for sugars when compared to the other ionic forms. However, when compared to calcium and sodium ones, resins in potassium cationic forms were shown to be the less efficient separating sugar mixtures. The resin with best separation performance was the Diaion UBK535Ca. A recovery yield of 92% (w/w) of FOS with 90% (w/w) of purity was obtained from batch experiments conducted in a single column loaded with the Diaion UBK535Ca resin at 25°C. The temperature shown did not influence the separation performance significantly. By increasing the column length, the purity of FOS increased to 92% (w/w), however the recovery yield decreased to 88% (w/w). Copyright © 2013 Elsevier B.V. All rights reserved.

  9. Strategies for the production of high-content fructo-oligosaccharides through the removal of small saccharides by co-culture or successive fermentation with yeast.

    PubMed

    Nobre, C; Castro, C C; Hantson, A-L; Teixeira, J A; De Weireld, G; Rodrigues, L R

    2016-01-20

    Fructo-oligosaccharides (FOS) obtained by fermentation of sucrose may be purified at large-scale by continuous chromatography (Simulated Moving Bed: SMB). In order to improve the efficiency of the subsequent SMB purification, the optimization of the fermentative broth composition in salts and sugars was investigated. Fermentations conducted at reduced amount of salts, using Aureobasidium pullulans whole cells, yielded 0.63 ± 0.03 g of FOS per gram of initial sucrose. Additionally, a microbial treatment was proposed to reduce the amount of small saccharides in the mixture. Two approaches were evaluated, namely a co-culture of A. pullulans with Saccharomyces cerevisiae; and a two-step fermentation in which FOS were first synthesized by A. pullulans and then the small saccharides were metabolized by S. cerevisiae. Assays were performed in 100mL shaken flasks and further scaled-up to a 3 L working volume bioreactor. Fermentations in two-step were found to be more efficient than the co-culture ones. FOS were obtained with a purity of 81.6 ± 0.8% (w/w), on a dry weight basis, after the second-step fermentation with S. cerevisiae. The sucrose amount was reduced from 13.5 to 5.4% in total sugars, which suggests that FOS from this culture broth will be more efficiently separated by SMB. Copyright © 2015 Elsevier Ltd. All rights reserved.

  10. Effect of fructo-oligosaccharide and isomalto-oligosaccharide addition on baking quality of frozen dough.

    PubMed

    Park, Eun Young; Jang, Sung-Bum; Lim, Seung-Taik

    2016-12-15

    The baking quality of frozen doughs containing different levels of fructo-oligosaccharides (FO) or isomalto-oligosaccharides (IMO) (3-9%, w/w flour), and stored for 0-8weeks at -18°C, was examined. The addition of FO or IMO increased the proof volume of the dough and the loaf volume of bread prepared from frozen dough. A 6% addition of FO or IMO was optimum, giving the highest proof volume and bread loaf volume, but a higher concentration than 6% induced low baking quality including lower proof volume and bread loaf volume. The bread crumb was moister and softer after the addition of FO or IMO before, and even after, frozen storage. Darker crumb colour was observed in the bread after the addition of FO or IMO. The oligosaccharides added to the frozen dough were effective in improving the quality of bread made from frozen dough, except for resulting in a darker bread crumb. Copyright © 2016. Published by Elsevier Ltd.

  11. Association of Lactobacillus crispatus with fructo-oligosaccharides and ascorbic acid in hydroxypropyl methylcellulose vaginal insert.

    PubMed

    Vitali, Beatrice; Abruzzo, Angela; Parolin, Carola; Palomino, Rogers Alberto Ñahui; Dalena, Francesco; Bigucci, Federica; Cerchiara, Teresa; Luppi, Barbara

    2016-01-20

    The aim of this work was to develop a synbiotic vaginal insert containing the probiotic strain Lactobacillus crispatus BC5, the prebiotic substrate fructo-oligosaccharide and the antioxidant agent ascorbic acid, for the prophylaxis and therapy of vaginal infections. Mucoadhesive in situ gelling vaginal inserts based on hydroxypropyl methylcellulose were prepared by freeze-drying, stored at +2-8 °C for 90 days and characterized in terms of technological and functional properties. Complete survival of L. crispatus BC5 was found immediately after insert preparation (96.08%) as well as after 90 days of storage (95.82%) in the vaginal inserts containing fructo-oligosaccharide, ascorbic acid and skimmed milk. Synbiotic inserts showed improved mucoadhesion ability (from three- to five-fold) with respect to a standard formulation based on hydroxypropyl methylcellulose alone. Moreover, inserts allowed to modulate lactobacilli release in virtue of the different amounts of fructo-oligosaccharide. Finally, antimicrobial activity was exerted by L. crispatus BC5 released from the vaginal formulation.

  12. Production of inulinase, fructosyltransferase and sucrase from fungi on low-value inulin-rich substrates and their use in generation of fructose and fructo-oligosaccharides.

    PubMed

    Rawat, Hemant Kumar; Ganaie, Mohd Anis; Kango, Naveen

    2015-03-01

    Owing to applications in the food and nutraceutical industries, inulinases, fructosyltransferases and sucrases have gained considerable attention in recent times. Twenty-five fungal strains were screened for production of these enzymes on three different media formulated using inulin-rich plant extracts prepared from asparagus root, dahlia tuber and dandelion root extract. Culture filtrates of the fungi were examined for hydrolytic activities. Fungi belonging to genus Aspergillus, A. niger GNCC 2655 (11.3 U/ml), A. awamori MTCC 2879 (8.2 U/ml), A. niger ATCC 26011 (7.9 U/ml) secreted high titers of inulinase followed by Penicillium sp. NFCCI 2768 (2.6 U/ml) and Penicillium citrinum MTCC 1256 (1.1 U/ml). High sucrase activity was noticed in A. niger GNCC 2613 (113 U/ml) and A. awamori MTCC 2879 (107.8 U/ml). Analysis of end products of inulinase action by HPLC revealed that most of the enzymes were exo-inulinases liberating fructose exclusively from inulin. Five fungi, P. citrinum MTCC 1256, Penicillium rugulosum MTCC 3487, Penicillium sp. NFCCI 2768, A. fumigatus GNCC 1351 and A. niger ATCC 26011 however, produced a mixture of endo- and exo-inulinases liberating oligosaccharides (GF3 and GF2) along with fructose. High inulinase/sucrase yielding strains were evaluated for extracellular and intracellular hydrolytic and transfructosylating activities and intracellular enzyme profiles were found to be considerably different in terms of titers and end products.

  13. Structural analyses and immunomodulatory properties of fructo-oligosaccharides from onion (Allium cepa).

    PubMed

    Kumar, V Prasanna; Prashanth, K V Harish; Venkatesh, Y P

    2015-03-06

    Onion (Allium cepa) is an immune-boosting food rich in fructans. The major aim of this study is to characterize and investigate the immunomodulatory properties of onion fructo-oligosaccharides (FOS). FOS was isolated from onion bulbs by hot 80% ethanol extraction (yield: ∼4.5 g/100 g fw) followed by gel permeation chromatography. NMR of onion FOS revealed unusual β-D-Glc terminal residue at the non-reducing end. TLC and ESI-MS analyses showed that onion FOS ranged from trisaccharides to hexasaccharides. Onion FOS (50 μg/mL) significantly increased (∼3-fold) the proliferation of mouse splenocytes/thymocytes vs. control. Further, onion FOS enhanced (∼2.5-fold) the production of nitric oxide by peritoneal exudates cells (PECs) from Wistar rats; intracellular free radicals production and phagocytic activity of isolated murine PECs were also augmented. Our structural and in vitro results indicate that onion FOS comprising of tri- to hexasaccharide units belongs to inulin-type fructans, and possess immunostimulatory activities towards murine lymphocytes and macrophages.

  14. Kinetics and model development for enzymatic synthesis of fructo-oligosaccharides using fructosyltransferase.

    PubMed

    Kashyap, Rahul; Palai, Tapas; Bhattacharya, Prashant K

    2015-12-01

    Experimental investigations were made to synthesize fructo-oligosaccharides (FOS) from sucrose using fructosyltransferase. The influence of various parameters such as temperature (45-55 °C), pH (4-5), initial sucrose concentration (ISC: 300-500 g/L) and enzyme concentration (4-32 U/mL) were varied. A maximum FOS yield of 60% was observed at ISC 500 g/L, pH 4.5 with enzyme activity 32 U/mL and at 55 °C. It was confirmed that 1-kestose (tri-) was the major product of FOS as compared to nystose (tetra-) and fructosylnystose (penta-saccharides). Further, the reaction rate increases with increase in temperature. From separate sets of experiments, it was observed that FOS formation was affected by glucose inhibition. Apart from the increase in the rate of FOS formation with increasing enzyme activity, the final values of FOS yield increase though till 16 U/mL and thereafter attain plateau. A kinetic model was also developed, based on Michaelis-Menten kinetics, and a five-step ten-parameter model, including glucose inhibition, was obtained. Model was solved using COPASI(®) (version 4.8) solver for kinetic parameter estimations followed by time course simulations.

  15. Randomised, double-blind, placebo-controlled trial of fructo-oligosaccharides in active Crohn's disease.

    PubMed

    Benjamin, Jane L; Hedin, Charlotte R H; Koutsoumpas, Andreas; Ng, Siew C; McCarthy, Neil E; Hart, Ailsa L; Kamm, Michael A; Sanderson, Jeremy D; Knight, Stella C; Forbes, Alastair; Stagg, Andrew J; Whelan, Kevin; Lindsay, James O

    2011-07-01

    The commensal intestinal microbiota drive the inflammation associated with Crohn's disease. However, bacteria such as bifidobacteria and Faecalibacterium prausnitzii appear to be immunoregulatory. In healthy subjects the intestinal microbiota are influenced by prebiotic carbohydrates such as fructo-oligosaccharides (FOS). Preliminary data suggest that FOS increase faecal bifidobacteria, induce immunoregulatory dendritic cell (DC) responses and reduce disease activity in patients with Crohn's disease. To assess the impact of FOS in patients with active Crohn's disease using an adequately powered randomised double-blind placebo-controlled trial with predefined clinical, microbiological and immunological end points. Patients with active Crohn's disease were randomised to 15 g/day FOS or non-prebiotic placebo for 4 weeks. The primary end point was clinical response at week 4 (fall in Crohn's Disease Activity Index of ≥ 70 points) in the intention-to-treat (ITT) population. 103 patients were randomised to receive FOS (n = 54) or placebo (n = 49). More patients receiving FOS (14 (26%) vs 4 (8%); p = 0.018) withdrew before the 4-week end point. There was no significant difference in the number of patients achieving a clinical response between the FOS and placebo groups in the ITT analysis (12 (22%) vs 19 (39%), p = 0.067). Patients receiving FOS had reduced proportions of interleukin (IL)-6-positive lamina propria DC and increased DC staining of IL-10 (p < 0.05) but no change in IL-12p40 production. There were no significant differences in the faecal concentration of bifidobacteria and F prausnitzii between the groups at baseline or after the 4-week intervention. An adequately powered placebo-controlled trial of FOS showed no clinical benefit in patients with active Crohn's disease, despite impacting on DC function. ISRCTN50422530.

  16. Absorption of calcium from milks enriched with fructo-oligosaccharides, caseinophosphopeptides, tricalcium phosphate, and milk solids.

    PubMed

    López-Huertas, Eduardo; Teucher, Birgit; Boza, Julio J; Martínez-Férez, Antonio; Majsak-Newman, Gosia; Baró, Luis; Carrero, Juan J; González-Santiago, María; Fonollá, Juristo; Fairweather-Tait, Susan

    2006-02-01

    Adequate intakes of calcium are required for optimal bone health and protection against chronic disease. Dairy products are an excellent source of calcium. The absorption of calcium from a range of fortified milks was measured in humans with the use of stable isotopes. Fifteen volunteers participated in a randomized, controlled, double-blind crossover study. Five types of semi-skimmed (1.9% fat) milk drinks were administered with a light breakfast: standard milk (control milk); milk enriched with calcium from milk solids and tricalcium phosphate [(TCP) MSS milk]; milk enriched with calcium from concentrated milk (CON milk); milk with added fructo-oligosaccharides [(FOSs) FOS milk]; and milk with added caseinophosphopeptides [(CPPs) CPP milk]. All the milks were labeled with 42Ca as CaCl2. The MSS milk was also labeled with 44Ca as TCP. The quantity of calcium in each drink was kept the same by varying the volume given. Calcium absorption did not differ significantly between the control milk and the calcium-fortified milks (MSS and CON milk) or the FOS and CPP milks. However, calcium absorption from the TCP added to the MSS milk was significantly higher than that from the control milk (27.5 +/- 7.6% and 24.5 +/- 7.3%, respectively; P = 0.003). Calcium-enriched milks are a valuable source of well-absorbed calcium. Absorption of added calcium as TCP was higher than that of calcium from the control milk, but the addition of FOSs or CPPs did not significantly increase calcium absorption. Further research is needed to ascertain the cost-effectiveness and public health benefits of consuming fortified milks.

  17. Structure of fructo-oligosaccharides from leaves and stem of Agave tequilana Weber, var. azul.

    PubMed

    Praznik, Werner; Löppert, Renate; Cruz Rubio, Josè M; Zangger, Klaus; Huber, Anton

    2013-11-15

    Fructo-oligosaccharides (FOSs) of a six year old agave plant variety, Agave tequilana, were isolated and fractionated by 2D preparative chromatography (SEC and rpHPLC). Structural analyses of different FOS-fractions were performed by reductive methylation analysis connected to GC/FID identification and NMR-analysis. FOSs from leaves (d.p. 3-8) contain single α-d-Glcp residues as well in terminal as internal position, however (2→1)-linked β-d-Fruf residues only. FOSs from stem, however, contain as well (2→1)- and (2→6)-linked β-d-Fruf residues with branched oligomeric repeating units. These characteristics indicate an enzymatically catalyzed metabolic regulation for the biosynthesis and transformation of fructans in A. tequilana which strongly depends on location and transport activities.

  18. Strategy for biotechnological process design applied to the enzymatic hydrolysis of agave fructo-oligosaccharides to obtain fructose-rich syrups.

    PubMed

    García-Aguirre, Mauricio; Sáenz-Alvaro, Victor A; Rodríguez-Soto, Mayra A; Vicente-Magueyal, Francisco J; Botello-Alvarez, Enrique; Jimenez-Islas, Hugo; Cárdenas-Manríquez, Marcela; Rico-Martínez, Ramiro; Navarrete-Bolaños, Jose L

    2009-11-11

    A strategy to optimize biotechnological process design is illustrated for the production of fructose-rich syrups via enzymatic hydrolysis of agave fructo-oligosaccharides. The optimization process includes ecological studies from natural fermentations leading to the selection of a strain with capacity for inulinase synthesis, and variable optimization for the synthesis, and enzymatic hydrolysis using the response surface methodology. The results lead to the selection of Kluyveromyces marxianus , endogenous strains isolated from aguamiel (natural fermented sugary sap from agave plants), as the main strain with high capacity for enzyme synthesis with inulinase activity. Production optimization at bioreactor level revealed that operation at 30.6 degrees C, 152 rpm, 1.3 VVM of aeration, and pH 6.3 leads to maximum inulinase synthesis, whereas 31 degrees C, 50 rpm, and pH 6.2 leads to maximum hydrolysis of agave fructo-oligosaccharides. HPLC analysis of the fructose-rich syrups obtained at these optimal conditions showed an average composition of 95% of fructose and 5% of glucose and the absence of sucrose. The analysis also revealed that the syrups are free of residues and toxic compounds, an undesirable occurrence often present when traditional methods based on thermal or acid hydrolysis are applied for their obtainment. Therefore, the product may be suitable for use as additive in many applications in the food and beverage industries.

  19. A fermented milk concentrate and a combination of short-chain galacto-oligosaccharides/long-chain fructo-oligosaccharides/pectin-derived acidic oligosaccharides protect suckling rats from rotavirus gastroenteritis.

    PubMed

    Rigo-Adrover, Mar; Pérez-Berezo, Teresa; Ramos-Romero, Sara; van Limpt, Kees; Knipping, Karen; Garssen, Johan; Knol, Jan; Franch, Àngels; Castell, Margarida; Pérez-Cano, Francisco J

    2017-01-01

    Human milk contains bioactive compounds that confer a protective role against gastrointestinal infections. In order to find supplements for an infant formula able to mimic these benefits of breast-feeding, two different concepts were tested. The products consisted of the following: (1) a Bifidobacterium breve- and Streptococcus thermophilus-fermented formula and (2) a combination of short-chain galacto-oligosaccharides/long-chain fructo-oligosaccharides with pectin-derived acidic oligosaccharides. A rotavirus infection suckling rat model was used to evaluate improvements in the infectious process and in the immune response of supplemented animals. Both nutritional concepts caused amelioration of the clinical symptoms, even though this was sometimes hidden by softer stool consistency in the supplemented groups. Both products also showed certain modulation of immune response, which seemed to be enhanced earlier and was accompanied by a faster resolution of the process. The viral shedding and the in vitro blocking assay suggest that these products are able to bind the viral particles, which can result in a milder infection. In conclusion, both concepts evaluated in this study showed interesting protective properties against rotavirus infection, which deserve to be investigated further.

  20. Prevention of coprophagy modifies magnesium absorption in rats fed with fructo-oligosaccharides.

    PubMed

    Ohta, A; Baba, S; Ohtsuki, M; Taguchi, A; Adachi, T

    1996-05-01

    We developed a new type of anal cup for prevention of coprophagy and determined whether the absorption of Ca and Mg and the stimulatory effects of feeding fructo-oligosaccharides (FO) on the absorption of Ca and Mg were altered by prevention of coprophagy in rats. Rats were fed on a FO-free diet or a diet containing 50 g FO/kg for 2 weeks with or without prevention of coprophagy. FO-feeding increased the apparent absorptive ratio of Ca and Mg in rats with or without prevention of coprophagy. However, in the FO-fed groups the absorptive ratio of Mg in rats with prevention of coprophagy was higher than in rats without prevention of coprophagy. The Ca content of the femur was higher in rats fed on the FO-diet than in rats fed on the FO-free diet both with and without coprophagy. In conclusion, FO-feeding increased the absorption of Ca and Mg in rats both with and without coprophagy. Moreover, prevention of coprophagy enhanced the absorption of Mg in rats fed with FO. Coprophagy has to be considered when the effects of luminal fermentation or mineral absorption are examined in rats.

  1. Fructo-oligosaccharide supplementation: effects on metabolic, endocrine and hematological traits in veal calves.

    PubMed

    Kaufhold, J; Hammon, H M; Blum, J W

    2000-02-01

    Fructo-oligosaccharides (FOS) are soluble fibres which exert various effects in the gastrointestinal tract, and induce metabolic and endocrine changes. The effects are favourable in diabetes mellitus, and may be favourable in veal calves, which during late periods of fattening often develop hyperglycemia, glucosuria and insulin resistance, especially during high lactose intake. Based on this we have studied metabolic, endocrine and haematological traits in veal calves (Simmental x Red Holstein) fed FOS (10 g/day; group GrF) or no FOS (group GrC). Whole milk and milk replacer in both groups, on a kg body weight basis, were fed in identical amounts. Experiments, lasting for 3 weeks, started when calves were 10 weeks old and weighed 117 kg. During week 3 lactose was supplemented to enhance post-absorptive glucose loads. Feed intakes were similar in both groups, but weight gain tended to be higher in GrF than GrC. The post-prandial increase of glucose concentrations was significantly smaller, of lactate tended to be smaller, and growth hormone peak frequency tended to be lower, whereas maximal insulin concentrations reached post-prandially were significantly higher in GrF than GrC. Eosinophil granulocytes increased during FOS feeding. In conclusion, FOS had basically similar effects on metabolic and endocrine traits in veal calves as in animals and humans with diabetes mellitus, but changes were small, albeit more prominent after lactose loads.

  2. Dietary supplementation with short-chain fructo-oligosaccharides improves insulin sensitivity in obese horses.

    PubMed

    Respondek, F; Myers, K; Smith, T L; Wagner, A; Geor, R J

    2011-01-01

    Obesity and insulin resistance are risk factors for laminitis in horses and ponies, and diet can play an important role in modulating these risk factors. Dietary supplementation with prebiotic fibers, such as short-chain fructo-oligosaccharides (scFOS), has resulted in improvement of insulin sensitivity in obese dogs and rodents. Thus, we hypothesized that scFOS may reduce insulin resistance in obese horses and designed a study to evaluate the effect of dietary supplementation with scFOS on insulin sensitivity. Eight mature Arabian geldings (BW = 523.0 ± 56.5 kg) with an average BCS of 8 were included in a crossover study. In each period, 4 horses were provided 45 g/d per horse of maltodextrin (control) and 4 horses received the same amount of scFOS for 6 wk, with a 3-wk washout between periods. Resting plasma concentrations of glucose, insulin, triglycerides, and leptin were measured. Minimal model analysis of a frequently sampled intravenous glucose tolerance test was used to evaluate insulin sensitivity, glucose effectiveness, acute insulin response to glucose, and disposition index. Without affecting BW and BCS, dietary supplementation with scFOS increased (P < 0.05) insulin sensitivity and reduced (P < 0.05) acute insulin response to glucose in comparison with maltodextrin but did not alter (P > 0.05) glucose effectiveness and disposition index. Resting serum insulin concentration also was reduced (P < 0.05) by scFOS supplementation but not by maltodextrin (P > 0.05). There was no effect (P > 0.05) of scFOS supplementation on plasma glucose or serum triglyceride and leptin concentrations. This study demonstrated that scFOS can moderately improve insulin sensitivity of obese horses, a finding that has potential relevance to the dietary management of obese, insulin-resistant horses at increased risk for laminitis.

  3. Dietary fructo-oligosaccharides and lactulose inhibit intestinal colonisation but stimulate translocation of salmonella in rats

    PubMed Central

    Bovee-Oudenhoven, I M J; ten Bruggencate, S J M; Lettink-Wissink, M L G; van der Meer, R

    2003-01-01

    Background and aims: It is frequently assumed that dietary non-digestible carbohydrates improve host resistance to intestinal infections by stimulating the protective gut microflora. However, compelling scientific evidence from in vivo infection studies is lacking. Therefore, we studied the effect of several non-digestible carbohydrates on the resistance of rats to Salmonella enteritidis infection. Methods: Rats (n=8 per group) were fed “humanised” purified diets containing 4% lactulose, fructo-oligosaccharides (FOS), resistant starch, wheat fibre, or cellulose. After an adaptation period of 2 weeks the animals were orally infected with S enteritidis. Supplement induced changes in faecal biochemical and microbiological parameters were studied before infection. Colonisation of salmonella was determined by studying the faecal excretion of this pathogen and translocation by analysis of urinary nitric oxide metabolites over time and classical organ cultures. Intestinal mucosal myeloperoxidase activity was determined to quantify intestinal inflammation after infection. Results: Despite stimulation of intestinal lactobacilli and bifidobacteria and inhibition of salmonella colonisation, FOS and lactulose significantly enhanced translocation of this pathogen. These supplements also increased cytotoxicity of faecal water and faecal mucin excretion, which may reflect mucosal irritation. In addition, caecal and colonic, but not ileal, mucosal myeloperoxidase activity was increased in infected rats fed FOS and lactulose. In contrast, cellulose, wheat fibre, and resistant starch did not affect the resistance to salmonella. Conclusions: In contrast to most expectations, FOS and lactulose impair the resistance of rats to intestinal salmonella infection. Obviously, stimulation of the endogenous lactobacilli and bifidobacteria is no guarantee of improved host defence against intestinal infections. PMID:14570725

  4. Effects of fructo-oligosaccharide-supplemented infant cereal: a double-blind, randomized trial.

    PubMed

    Moore, Nancy; Chao, Cewin; Yang, Li-Ping; Storm, Heidi; Oliva-Hemker, Maria; Saavedra, Jose M

    2003-09-01

    Fructo-oligosaccharides (FOS) may have potential benefits, since they exhibit many soluble dietary fibre-like properties. Foods currently available for weaning infants are generally low in fibre content and lack these potential benefits. Data documenting tolerance of FOS in weaning foods are greatly lacking. Our present objective was to evaluate the tolerance and gastrointestinal effects of FOS-supplemented infant cereal used as a daily addition to the diet of healthy infants. Healthy infants were randomly assigned to receive either 0.75 g FOS per serving of cereal or placebo for 28 d. The primary outcome of interest was gastrointestinal tolerance, which was assessed by daily parental reporting of functional variables for 28 d, including stool patterns and signs and symptoms of gastrointestinal tolerance. Secondary outcomes were also measured including: cereal intake (g cereal and g FOS/d), stool pH, changes in anthropometric measurements and adverse events. The study population included a total of fifty-six infants, age range 16.2-46.2 weeks with a mean age of 32.5 (sd 8.9) weeks; twenty-nine infants were randomized to the control group (age 31.8 (sd 9.0) weeks) and twenty-seven to the FOS-supplemented group (34.7 (sd 8.9) weeks). Average daily total intake per infant and average intake per serving were similar in both groups. Average FOS consumption was 0.74 (sd 0.39) g/d and as high as 3.00 g/d. Stool consistency was less likely to be described as 'hard', and more likely to be described as 'soft' or 'loose', in the FOS v. control group. The mean number of stools per infant was 1.99 (sd 0.62) per d in the FOS-supplemented group compared with 1.58 (sd 0.66) in the control group (P=0.02). There were no differences between the groups in reporting for crying, spitting-up or colic. No differences were found for stool pH. FOS-supplements added to cereal were well tolerated in doses of up to 3.00 g/d. FOS consumption led to more regular and softer stools, without

  5. Chemical characterization and prebiotic activity of fructo-oligosaccharides from Stevia rebaudiana (Bertoni) roots and in vitro adventitious root cultures.

    PubMed

    Sanches Lopes, Sheila Mara; Francisco, Mariane Grigio; Higashi, Bruna; de Almeida, Rafaela Takako Ribeiro; Krausová, Gabriela; Pilau, Eduardo Jorge; Gonçalves, José Eduardo; Gonçalves, Regina Aparecida Correia; Oliveira, Arildo José Braz de

    2016-11-05

    Stevia rebaudiana (Bertoni) is widely studied because of its foliar steviol glycosides. Fructan-type polysaccharides were recently isolated from its roots. Fructans are reserve carbohydrates that have important positive health effects and technological applications in the food industry. The objective of the present study was to isolate and characterize fructo-oligosaccharides (FOSs) from S. rebaudiana roots and in vitro adventitious root cultures and evaluate the potential prebiotic effect of these molecules. The in vitro adventitious root cultures were obtained using a roller bottle system. Chemical analyses (gas chromatography-mass spectrometry, (1)H nuclear magnetic resonance, and off-line electrospray ionization-mass spectrometry) revealed similar chemical properties of FOSs that were obtained from the different sources. The potential prebiotic effects of FOSs that were isolated from S. rebaudiana roots enhanced the growth of both bifidobacteria and lactobacilli, with strains specificity in their fermentation ability.

  6. A new natural source for obtainment of inulin and fructo-oligosaccharides from industrial waste of Stevia rebaudiana Bertoni.

    PubMed

    Lopes, Sheila Mara Sanches; Krausová, Gabriela; Carneiro, José Walter Pedroza; Gonçalves, José Eduardo; Gonçalves, Regina Aparecida Correia; de Oliveira, Arildo José Braz

    2017-06-15

    Fructan-type inulin and fructo-oligosaccharides (FOS) are reserve polysaccharides that offer an interesting combination of nutritional and technological properties for food industry. Stevia rebaudiana is used commercially in the sweetener industry due to the high content of steviol glycosides in its leaves. With the proposal of using industrial waste, the objective of the present study was to isolate, characterize and evaluate the prebiotic activity of inulin and FOS from S. rebaudiana stems. The chemical characterization of the samples by GC-MS, NMR and off-line ESI-MS showed that it was possible to obtain inulin molecules from the S. rebaudiana stems with a degree of polymerization (DP) of 12, and FOS with a DP<6. The in vitro prebiotic assay of these molecules indicates a strain specificity in fermentation capacity of fructans as substrate. FOS molecules with a low DP are preferably fermented by beneficial microbiota than inulin molecules with higher DP.

  7. Effect of oligosaccharides on the growth of Lactobacillus delbrueckii subsp. bulgaricus strains isolated from dairy products.

    PubMed

    Ignatova, Tseteslava; Iliev, Ilia; Kirilov, Nikolai; Vassileva, Tonka; Dalgalarrondo, Michèle; Haertlé, Thomas; Chobert, Jean-Marc; Ivanova, Iskra

    2009-10-28

    Eighteen lactic acid bacteria (LAB) strains isolated from dairy products, all identified as Lactobacillus delbrueckii subsp. bulgaricus, were tested for their ability to grow on three different oligosaccharides: fructo-oligosaccharides (FOS), gluco-oligosaccharides (GOS) and galacto-oligosaccharides (GalOS). The growth of LAB on different oligosaccharides was very different. Study of the antimicrobial activities of these LAB indicated that the system of uptake of unusual sugars influenced in a specific way the production of antimicrobial substances (bacteriocins) specific against gram-negative bacteria. The added oligosaccharides induced LAB to form end-products of a typical mixed acid fermentation. The utilization of different types of oligosaccharides may help to explain the ability of Lactobacillus strains to compete with other bacteria in the ecosystem of the human gastro-intestinal tract.

  8. Chemically Defined Diet Alters the Protective Properties of Fructo-Oligosaccharides and Isomalto-Oligosaccharides in HLA-B27 Transgenic Rats

    PubMed Central

    Valcheva, Rosica; Gänzle, Michael G.; Dieleman, Levinus A.

    2014-01-01

    Non-digestible oligosaccharides (NDO) were shown to reduce inflammation in experimental colitis, but it remains unclear whether microbiota changes mediate their colitis-modulating effects. This study assessed intestinal microbiota and intestinal inflammation after feeding chemically defined AIN-76A or rat chow diets, with or without supplementation with 8 g/kg body weight of fructo-oligosaccharides (FOS) or isomalto-oligosaccharides (IMO). The study used HLA-B27 transgenic rats, a validated model of inflammatory bowel disease (IBD), in a factorial design with 6 treatment groups. Intestinal inflammation and intestinal microbiota were analysed after 12 weeks of treatment. FOS and IMO reduced colitis in animals fed rat chow, but exhibited no anti-inflammatory effect when added to AIN-76A diets. Both NDO induced specific but divergent microbiota changes. Bifidobacteria and Enterobacteriaceae were stimulated by FOS, whereas copy numbers of Clostridium cluster IV were decreased. In addition, higher concentrations of total short-chain fatty acids (SCFA) were observed in cecal contents of rats on rat chow compared to the chemically defined diet. AIN-76A increased the relative proportions of propionate, iso-butyrate, valerate and iso-valerate irrespective of the oligosaccharide treatment. The SCFA composition, particularly the relative concentration of iso-butyrate, valerate and iso-valerate, was associated (P≤0.004 and r≥0.4) with increased colitis and IL-1 β concentration of the cecal mucosa. This study demonstrated that the protective effects of fibres on colitis development depend on the diet. Although diets modified specific cecal microbiota, our study indicates that these changes were not associated with colitis reduction. Intestinal inflammation was positively correlated to protein fermentation and negatively correlated with carbohydrate fermentation in the large intestine. PMID:25369019

  9. Influence of a blend of fructo-oligosaccharides and sugar beet fiber on nutrient digestibility and plasma metabolite concentrations in healthy beagles.

    PubMed

    Diez, M; Hornick, J L; Baldwin, P; Istasse, L

    1997-11-01

    To evaluate effects of a blend of fructo-oligosaccharides and sugar beet fiber (4:1) at 3 incorporation rates on nutrient digestibility and plasma glucose, insulin, alpha-aminonitrogen, urea, cholesterol, and triglycerides concentrations measured weekly in nonfed dogs and during a 360-minute period after a meal. 8 castrated 1- to 1.4-year-old young adult male Beagles weighing 10.0 to 13.5 kg. Diets containing 2 incorporation rates of a blend of fructo-oligosaccharides and sugar beet fiber (5 and 10% on a dry matter basis [diets B and C, respectively]) were compared with a control diet without additional fiber (diet A). The 3 diets were evaluated for ability to modify digestibility of dry and organic matter, protein, fat, and ash and for effects on plasma glucose, insulin, alpha-aminonitrogen, urea, cholesterol, and triglycerides concentrations. Each diet was fed for 6 weeks; plasma samples were collected weekly before feeding and after feeding on the last day of the period. During 1 week at the end of the 6-week period, dogs were kept in metabolic cages. Each period of the block was followed by a 4-week washout period. Incorporating the blend of fructo-oligosaccharides and sugar beet fiber in the diet was associated with greater passage of wet feces (diets B and C) and lower protein digestibility (diet C). Postprandial glucose (diet C), urea (diets B and C) and triglyceride (diets B and C) concentrations were significantly (P < 0.01) decreased. Weekly preprandial measurements were characterized by decreased urea (diets B and C), cholesterol (diet C), and triglycerides (diets B and C) concentrations (P < 0.001). Chronic consumption of fermentable fiber is associated with mildly decreased protein digestibility and with metabolic effects in nonfed or fed dogs. A blend of fructo-oligosaccharides and sugar beet fiber should be tested as a dietary aid for treatment of chronic diseases, such as diabetes mellitus or hyperlipidemia, in dogs.

  10. Presence of Inulin-Type Fructo-Oligosaccharides and Shift from Raffinose Family Oligosaccharide to Fructan Metabolism in Leaves of Boxtree (Buxus sempervirens)

    PubMed Central

    Van den Ende, Wim; Coopman, Marlies; Vergauwen, Rudy; Van Laere, André

    2016-01-01

    Fructans are known to occur in 15% of flowering plants and their accumulation is often associated with stress responses. Typically, particular fructan types occur within particular plant families. The family of the Buxaceae, harboring Pachysandra terminalis, an accumulator of graminan- and levan-type fructans, also harbors boxtree (Buxus sempervirens), a cold and drought tolerant species. Surprisingly, boxtree leaves do not accumulate the expected graminan- and levan-type fructans, but small inulin fructo-oligosaccharides (FOS: 1-kestotriose and nystose) and raffinose family oligosaccharides (RFOs: raffinose and stachyose) instead. The seasonal variation in concentrations of glucose, fructose, sucrose, FOS and RFOs were followed. Raffinose and stachyose peaked during the winter months, while FOS peaked at a very narrow time-interval in spring, immediately preceded by a prominent sucrose accumulation. Sucrose may function as a reserve carbohydrate in winter and early spring leaves. The switch from RFO to fructan metabolism in spring strongly suggests that fructans and RFOs fulfill distinct roles in boxtree leaves. RFOs may play a key role in the cold acclimation of winter leaves while temporal fructan biosynthesis in spring might increase sink strength to sustain the formation of new shoots. PMID:26973663

  11. Gut morphology and hepatic oxidative status of European sea bass (Dicentrarchus labrax) juveniles fed plant feedstuffs or fishmeal-based diets supplemented with short-chain fructo-oligosaccharides and xylo-oligosaccharides.

    PubMed

    Guerreiro, Inês; Couto, Ana; Pérez-Jiménez, Amalia; Oliva-Teles, Aires; Enes, Paula

    2015-12-28

    The effects of short-chain fructo-oligosaccharides (scFOS) and xylo-oligosaccharides (XOS) on gut morphology and hepatic oxidative status were studied in European sea bass juveniles weighing 60 g. Fish were fed diets including fishmeal (FM diets) or plant feedstuffs (PF diets; 30 FM:70 PF) as main protein sources (control diets). Four other diets were formulated similar to the control diets but including 1 % scFOS or 1 % XOS. At the end of the trial, fish fed PF-based diets presented histomorphological alterations in the distal intestine, whereas only transient alterations were observed in the pyloric caeca. Comparatively to fish fed FM-based diets, fish fed PF diets had higher liver lipid peroxidation (LPO), superoxide dismutase (SOD) and catalase (CAT) activities, and lower glutathione peroxidase, glutathione reductase and glucose 6-phosphate dehydrogenase activities. In fish fed the PF diets, prebiotic supplementation decreased SOD activity and XOS supplementation further decreased CAT activity. In fish fed the FM diets, XOS supplementation promoted a reduction of all antioxidant enzyme activities. Overall, dietary XOS and scFOS supplementation had only minor effects on gut morphology or LPO levels. However, dietary XOS reduced antioxidant enzymatic activity in both PF and FM diets, which indicate a positive effect on reduction of hepatic reactive oxygen species production.

  12. Transfer of blood urea nitrogen to cecal microbial nitrogen is increased by fructo-oligosaccharide feeding in guinea pigs.

    PubMed

    Kawasaki, Kiyonori; Min, Xiao; Li, Xiao; Hasegawa, Ena; Sakaguchi, Ei

    2015-01-01

    The present study was conducted to determine the mechanism by which nitrogen (N) availability is improved by fructo-oligosaccharide (FOS) in guinea pigs. Adult male guinea pigs were fed a commercial pellet diet (50 g/day) with either 5% glucose or 5% FOS for 7 days in individual metabolism cages. After 7 days of feeding the diet, (15) N-urea was administered intravenously 1 h before slaughter under anesthesia. The amount and concentration of total, protein, bacterial, ammonia and urea N and the (15) N atom % excess were measured in blood, liver, gut contents and urine. The (15) N atom % excess of total and protein N, and the amount of total, protein and bacteria N and (15) N in the cecum were significantly increased by the consumption of FOS. Furthermore, the concentration and amount of short-chain fatty acids were significantly increased by the consumption of FOS. In contrast, the amount of urinary (15) N was significantly decreased by the consumption of FOS. These results suggest that consumption of FOS increases transfer of blood urea N into the large intestine for bacterial N synthesis, which is subsequently re-absorbed by cecotrophy, and contributes to the increase of N utilization in guinea pigs.

  13. The effect of inulin and fructo-oligosaccharide supplementation on the textural, rheological and sensory properties of bread and their role in weight management: a review.

    PubMed

    Morris, Cécile; Morris, Gordon A

    2012-07-15

    There is evidence that fructo-oligosaccharides (FOS) and inulin can impart a range of health benefits if consumed on a regular basis. The health benefits include increased mineral absorption and improved immune response and while there is mounting evidence that prebiotics play a role in colorectal cancer prevention, their role of satiety and weight management is still being investigated. In this review we look at the evidence published so far on FOS or inulin supplementation and weight management. We also establish whether prebiotic enriched breads are feasible in terms of dough machinability, bread characteristics and consumers acceptance. Addition of inulin to bread generally resulted in smaller loaves with a harder crumb and darker colour. The limited sensory studies on those products reflect those findings and acceptability decreased with inulin content. However, a fortification of 5% seems achievable. Despite evidence that yeast invertase and dry heat degrade inulin, the extent to which this is the case and whether the prebiotics maintain their activity is not known. There is still a great deal of work to be done to establish whether a bread prepared with enough inulin to retain a significant activity can be manufactured without compromising consumer acceptance.

  14. Inulin and fructo-oligosaccharides have divergent effects on colitis and commensal microbiota in HLA-B27 transgenic rats.

    PubMed

    Koleva, Petya T; Valcheva, Rosica S; Sun, Xu; Gänzle, Michael G; Dieleman, Levinus A

    2012-11-14

    Modulation of intestinal microbiota by non-digestible carbohydrates may reduce inflammation in inflammatory bowel disease (IBD). The aim of the present study was to assess the effects of inulin and fructo-oligosaccharides (FOS) on intestinal microbiota and colitis in HLA-B27 transgenic rats, a well-validated rodent model for IBD. In this study, 4-week-old rats were fed 8 g/kg body weight inulin or FOS for 12 weeks, or not. Faeces were collected at 4 and 16 weeks of age; and caecal samples were collected at necropsy. The effects of inulin and FOS on chronic intestinal inflammation were assessed using a gross gut score, histology score and levels of mucosal IL-1β. Intestinal microbiota were characterised by quantitative PCR and denaturing gradient gel electrophoresis. Colitis was significantly reduced in all FOS-fed rats compared to the control diet, whereas inulin decreased chronic intestinal inflammation in only half the number of animals. Quantitative analysis of caecal microbiota demonstrated that inulin increased the numbers of total bacteria and the Bacteroides-Prevotella-Porphyromonas group, FOS increased bifidobacteria, and both fructans decreased Clostridium cluster XI. In the faecal samples, both inulin and FOS decreased total bacteria, Bacteroides-Prevotella-Porphyromonas group, and Clostridium clusters XI and XIVa. FOS increased Bifidobacterium spp., and mediated a decrease of gene copies of Enterobacteriaceae and Clostridium difficile toxin B in faeces. SCFA concentrations in the faecal and caecal samples were unaffected by the diets. In conclusion, FOS increased the abundance of Bifidobacterium spp., whereas both fructans reduced Clostridium cluster XI and C. difficile toxin gene expression, correlating with a reduction of chronic intestinal inflammation.

  15. Lactobacillus fermentum BR11 and fructo-oligosaccharide partially reduce jejunal inflammation in a model of intestinal mucositis in rats.

    PubMed

    Smith, Cassie L; Geier, Mark S; Yazbeck, Roger; Torres, Diana M; Butler, Ross N; Howarth, Gordon S

    2008-01-01

    Although probiotics are beginning to enter mainstream medicine for disorders of the colon, their effects on the small bowel remain largely unexplored. We investigated the recently identified probiotic, Lactobacillus fermentum (L. fermentum) BR11 (BR11) and the prebiotic, fructo-oligosaccharide (FOS), both individually and in synbiotic combination, for their potential to alleviate intestinal mucositis. From Days 0-9, rats consumed skim milk (SM; saline + SM), low dose (LD-BR11; 1 x 10(6)cfu/ml), high dose (HD-BR11; 1 x 10(9)cfu/ml), LD-FOS (3%), HD-FOS (6%), or synbiotic (HD-BR11/FOS). On Day 7, rats were injected with 5-fluorouracil (5-FU; 150 mg/kg). All rats were sacrificed on Day 10. Intestinal tissues were collected for quantitative histology, sucrase, and myeloperoxidase (MPO) determinations. 5-FU decreased sucrase activity, villus height, crypt depth, and crypt cell proliferation compared to controls. Compared to 5-FU + SM, histological damage severity scores were increased for all treatments, although all were effective at reducing jejunal inflammation, indicated by reduced MPO activity (P < 0.05). The combination of BR11 and FOS did not provide additional protection. Moreover, HD-FOS and the synbiotic actually increased clinical mucositis severity (P < 0.05). We conclude that L. fermentum BR11 has the potential to reduce inflammation of the upper small intestine. However, its combination with FOS does not appear to confer any further therapeutic benefit for the alleviation of mucositis.

  16. Short-Chain Fructo-Oligosaccharides Modulate Intestinal Microbiota and Metabolic Parameters of Humanized Gnotobiotic Diet Induced Obesity Mice

    PubMed Central

    Respondek, Frederique; Gerard, Philippe; Bossis, Mathilde; Boschat, Laura; Bruneau, Aurélia; Rabot, Sylvie; Wagner, Anne; Martin, Jean-Charles

    2013-01-01

    Prebiotic fibres like short-chain fructo-oligosaccharides (scFOS) are known to selectively modulate the composition of the intestinal microbiota and especially to stimulate Bifidobacteria. In parallel, the involvement of intestinal microbiota in host metabolic regulation has been recently highlighted. The objective of the study was to evaluate the effect of scFOS on the composition of the faecal microbiota and on metabolic parameters in an animal model of diet-induced obesity harbouring a human-type microbiota. Forty eight axenic C57BL/6J mice were inoculated with a sample of faecal human microbiota and randomly assigned to one of 3 diets for 7 weeks: a control diet, a high fat diet (HF, 60% of energy derived from fat)) or an isocaloric HF diet containing 10% of scFOS (HF-scFOS). Mice fed with the two HF gained at least 21% more weight than mice from the control group. Addition of scFOS partially abolished the deposition of fat mass but significantly increased the weight of the caecum. The analysis of the taxonomic composition of the faecal microbiota by FISH technique revealed that the addition of scFOS induced a significant increase of faecal Bifidobacteria and the Clostridium coccoides group whereas it decreased the Clostridium leptum group. In addition to modifying the composition of the faecal microbiota, scFOS most prominently affected the faecal metabolome (e.g. bile acids derivatives, hydroxyl monoenoic fatty acids) as well as urine, plasma hydrophilic and plasma lipid metabolomes. The increase in C. coccoides and the decrease in C. leptum, were highly correlated to these metabolic changes, including insulinaemia, as well as to the weight of the caecum (empty and full) but not the increase in Bifidobacteria. In conclusion scFOS induce profound metabolic changes by modulating the composition and the activity of the intestinal microbiota, that may partly explain their effect on the reduction of insulinaemia. PMID:23951074

  17. Effect of Bifidobacterium thermophilum RBL67 and fructo-oligosaccharides on the gut microbiota in Göttingen minipigs.

    PubMed

    Tanner, Sabine A; Lacroix, Christophe; Del'Homme, Christophe; Jans, Christoph; Zihler Berner, Annina; Bernalier-Donadille, Annick; Chassard, Christophe

    2015-09-14

    Modulating the gut microbiota via dietary interventions is a common strategy to enhance the natural defence mechanisms of the host. Several in vitro studies have highlighted the probiotic potential of Bifidobacterium thermophilum RBL67 (RBL67) selected for its anti-Salmonella effects. The present study aimed to investigate the impact of RBL67 alone and combined with fructo-oligosaccharides (FOS) on the gut microbiota of Göttingen minipigs. Minipigs were fed a basal diet supplemented with 8 g/d probiotic powder (1×109 CFU/g in skim milk matrix) (probiotic diet (PRO)), 8 g/d probiotic powder plus 8 g/d FOS (synbiotic diet (SYN)) or 8 g/d skim milk powder (control), following a cross-sectional study design. Faecal and caecal microbiota compositions were analysed with pyrosequencing of 16S rRNA genes and quantitative PCR. Metabolic activity in the caecum and colon was measured by HPLC. 16S rRNA gene amplicon sequencing revealed that minipig faeces show close similarity to pig microbiota. During the treatments and at the time of killing of animals, RBL67 was consistently detected in faeces, caecum and colon at numbers of 105-106 16S rRNA copies/g content after feeding PRO and SYN diets. At the time of killing of animals, significantly higher Bifidobacterium numbers in the caecum and colon of SYN-fed minipigs were measured compared with PRO. Our data indicate that the Göttingen minipig may be a suitable model for gut microbiota research in pigs. Data from this first in vivo study of RBL67 colonisation suggest that the combination with FOS may represent a valuable symbiotic strategy to increase probiotic bacteria levels and survival in gastrointestinal tracts for feed and food applications.

  18. Short-chain fructo-oligosaccharides modulate intestinal microbiota and metabolic parameters of humanized gnotobiotic diet induced obesity mice.

    PubMed

    Respondek, Frederique; Gerard, Philippe; Bossis, Mathilde; Boschat, Laura; Bruneau, Aurélia; Rabot, Sylvie; Wagner, Anne; Martin, Jean-Charles

    2013-01-01

    Prebiotic fibres like short-chain fructo-oligosaccharides (scFOS) are known to selectively modulate the composition of the intestinal microbiota and especially to stimulate Bifidobacteria. In parallel, the involvement of intestinal microbiota in host metabolic regulation has been recently highlighted. The objective of the study was to evaluate the effect of scFOS on the composition of the faecal microbiota and on metabolic parameters in an animal model of diet-induced obesity harbouring a human-type microbiota. Forty eight axenic C57BL/6J mice were inoculated with a sample of faecal human microbiota and randomly assigned to one of 3 diets for 7 weeks: a control diet, a high fat diet (HF, 60% of energy derived from fat)) or an isocaloric HF diet containing 10% of scFOS (HF-scFOS). Mice fed with the two HF gained at least 21% more weight than mice from the control group. Addition of scFOS partially abolished the deposition of fat mass but significantly increased the weight of the caecum. The analysis of the taxonomic composition of the faecal microbiota by FISH technique revealed that the addition of scFOS induced a significant increase of faecal Bifidobacteria and the Clostridium coccoides group whereas it decreased the Clostridium leptum group. In addition to modifying the composition of the faecal microbiota, scFOS most prominently affected the faecal metabolome (e.g. bile acids derivatives, hydroxyl monoenoic fatty acids) as well as urine, plasma hydrophilic and plasma lipid metabolomes. The increase in C. coccoides and the decrease in C. leptum, were highly correlated to these metabolic changes, including insulinaemia, as well as to the weight of the caecum (empty and full) but not the increase in Bifidobacteria. In conclusion scFOS induce profound metabolic changes by modulating the composition and the activity of the intestinal microbiota, that may partly explain their effect on the reduction of insulinaemia.

  19. Study of the effect exerted by fructo-oligosaccharides from yacon (Smallanthus sonchifolius) root flour in an intestinal infection model with Salmonella Typhimurium.

    PubMed

    Velez, Eva; Castillo, Natalia; Mesón, Oscar; Grau, Alfredo; Bibas Bonet, María E; Perdigón, Gabriela

    2013-06-01

    Beneficial effects of prebiotics like inulin and fructo-oligosaccharides (FOS) have been proven in health and nutrition. Yacon (Smallanthus sonchifolius), an Andean crop, contains FOS (50–70% of its dry weight) and, therefore, is considered a prebiotic. Commercial FOS can upregulate total secretory IgA (S-IgA) in infant mice, prevent infection with Salmonella in swine or enhance immune response for Salmonella vaccine in a mouse model. Previously, we found that administration of yacon root flour regulates gut microbiota balance and has immunomodulatory effects without inflammatory responses. The aim of the present paper is to analyse if yacon prevents enteric infection caused by a strain of Salmonella enteritidis serovar Typhimurium (S. Typhimurium) in a mouse model. BALB/c mice were supplemented with yacon flour (45 d), challenged with S. Typhimurium and killed to study pathogen translocation, total and specific IgA production by ELISA, presence of IgA and other cytokines and Toll-like receptor 4 (TLR4) and clustor of differentiation 206 (CD206) receptors positive cells by immunofluorescence and histological changes. Yacon flour administration had a protective effect from 15 to 30 d of treatment. We found a peak of total S-IgA production without translocation of the pathogen for these periods. At 30 d, there was an increase in IL-6 and macrophage inflammatory proteins-1aþ cells and expression of the receptors CD206 and TLR4. Yacon flour did not have incidence in pathogen-specific S-IgA production. Longer periods (45 d) of administration had no protective effect. Therefore, yacon can prevent enteric infection caused by S. Typhimurium when given up to 30 d; this effect would be mediated by enhancing non-specific immunity, such as total S-IgA, that improves the immunological intestinal barrier.

  20. [Determination of fructo-oligosaccharides in milk powder by high performance liquid chromatography-quadrupole/electrostatic field orbitrap high resolution mass spectrometry].

    PubMed

    Liu, Yun; Ding, Tao; Xu, Suli; Wu, Bin; Shen, Chongyu; Zhang, Rui; Wang, Yan; Fei, Xiaoqing

    2015-10-01

    A method of high performance liquid chromatography-quadrupole/electrostatic field Orbitrap high resolution mass spectrometry (HPLC-Q/Orbitrap MS) was developed to determine fructo-oligosaccharides in milk powder. The milk powder samples were dissolved in deionized water. Subsequently, an aqueous solution of zinc acetate was used to precipitate protein. After centrifugation, the final aqueous solution was filtered by a polytetrafluoroethylene (PTFE) membrane with pore size of 0.22 μm. The analytes were separated on a Carbohydrate column (100 mm x 2.1 mm, 2.6 μm) through gradient elution with the combination of acetonitrile and 0.1% formic acid aqueous solution. The target-MS/MS templates were performed at isolation window of m/z 4.0 and collision energy of 30 eV in positive mode to extract the accurate product ion mass of analytes. Under the optimal condition, 1-kestose (GF2), nystose (GF3) and 1-F-β-fructofuranosyl nystose (GF4) were well separated and the accuracy of extracted mass routinely detected was below 5 x 10(-6) (5 ppm). The whole analysis time is only ten minutes. The detection limits for GF2 and GF3 were 100 μg/kg, and the detection limit for GF4 was 55 μg/kg. Good linearities were obtained in their respective linear ranges with correlation coefficients higher than 0.998. The average recoveries at three spiked levels (5, 10 and 20 mg/kg) were in the range of 75.8%-107.3% and the relative standard deviations (RSDs) were in the range of 1.6% - 8.3%. The proposed method is simple, sensitive, fast and only in need of precipitation of proteins. The interference of matrix can be eliminated through the selection of product ion. The results were convenient and reliable and thus can be used in the large batch determination of any milk powder.

  1. Maternal short-chain fructo-oligosaccharide supplementation increases intestinal cytokine secretion, goblet cell number, butyrate concentration and Lawsonia intracellularis humoral vaccine response in weaned pigs.

    PubMed

    Le Bourgot, Cindy; Le Normand, Laurence; Formal, Michèle; Respondek, Frédérique; Blat, Sophie; Apper, Emmanuelle; Ferret-Bernard, Stéphanie; Le Huërou-Luron, Isabelle

    2017-01-01

    Prebiotic supplementation modulates immune system development and function. However, less is known about the effects of maternal prebiotic consumption on offspring intestinal defences and immune system responsiveness. We investigated the effects of maternal short-chain fructo-oligosaccharide (scFOS) supplementation on mucin-secreting cells, ileal secretory IgA and cytokine secretion of weaned offspring and their humoral response to an oral vaccine against obligate intracellular Lawsonia intracellularis. Sows were fed a control diet (CTRL) or scFOS-supplemented diet during the last third of gestation and throughout lactation. At weaning, each litter was divided into two groups receiving a post-weaning CTRL or scFOS diet for a month. Pigs from the four groups were either non-vaccinated (n 16) or vaccinated (n 117) at day 33. Biomarkers related to intestinal defences and immune parameters were analysed 3 weeks later. SCFA production was assessed over time in suckling and weaned pigs. Maternal scFOS supplementation improved ileal cytokine secretions (interferon (IFN)-γ, P<0·05; IL-4, P=0·07) and tended to increase caecal goblet cell number (P=0·06). It increased IgA vaccine response in the serum (P<0·01) and ileal mucosa (P=0·08). Higher bacterial fermentative activity was observed during lactation (total faecal SCFA, P<0·001) and after weaning (colonic butyrate, P=0·10) in pigs from scFOS-supplemented mothers. No synergistic effect between maternal and post-weaning scFOS supplementation was observed. Therefore, maternal scFOS supplementation has long-lasting consequences by strengthening gut defences and immune response to a vaccine against an intestinal obligate intracellular pathogen. Prebiotic consumption by gestating and lactating mothers is decisive in modulating offspring intestinal immunity.

  2. Postprandial glycaemic and insulinaemic responses in adults after consumption of dairy desserts and pound cakes containing short-chain fructo-oligosaccharides used to replace sugars.

    PubMed

    Lecerf, J M; Clerc, E; Jaruga, A; Wagner, A; Respondek, F

    2015-01-01

    The present studies aimed to evaluate the glycaemic and insulinaemic responses, in healthy adults, to short-chain fructo-oligosaccharides (scFOS) from sucrose used to replace sugars in foods. Two study populations aged 18-50 years were recruited and they consumed dairy desserts or pound cakes containing either standard sugar content or scFOS to replace 30 % of the sugar content. For each study, the two products were tested once under a double-blind and cross-over design with at least 7 d between the two tests. Glucose and insulin were measured using standard methods in blood samples collected with a venous catheter for 120 min during a kinetic test. For the dairy desserts, replacing 30 % of the sugars with scFOS significantly reduced postprandial glycaemic (AUC0-120 min; P = 0·020) and insulinaemic (AUC0-120 min; P = 0·003) responses. For the pound cakes, the glycaemic response was not altered (AUC0-120 min; P =  0·322) while the insulinaemic response tended to be lower (AUC0-120 min; P = 0·067). This study showed that scFOS can be used to replace sugars with the benefit of lowering the postprandial glycaemic response without increasing the insulinaemic response. The effect might be modulated by other parameters (e.g. fat content) of the food matrices.

  3. Effect of sucrose concentration on the composition of enzymatically synthesized short-chain fructo-oligosaccharides as determined by FTIR and multivariate analysis.

    PubMed

    Romano, Nelson; Santos, Mauricio; Mobili, Pablo; Vega, Roberto; Gómez-Zavaglia, Andrea

    2016-07-01

    Fructo-oligosaccharides (FOS) are mixtures of oligosaccharides composed of fructose and glucose units. As their composition is determined by the synthesis conditions, the goals of this work were: (a) to engineer FOS of different composition by adjusting the sucrose concentration used as initial substrate; (b) to define partial least square (PLS) based-models to quantify all the sugars present in the reaction medium directly from the FTIR spectra. The yield of each reaction was calculated as the percentage of initial sucrose converted to each oligosaccharide, as monitored by HPLC. In parallel, the reactions were followed by FTIR. Six different PLS models aiming to determine the concentration of each carbohydrate present in the reaction medium were calibrated and independently validated. The means of predicted values fitted well to those obtained by HPLC. Determining FOS composition directly from the FTIR spectra represents a useful tool to monitor enzymatic synthesis, with strong impact at both an academic and an industrial level. Copyright © 2016 Elsevier Ltd. All rights reserved.

  4. Effects of short-chain fructo-oligosaccharides on glucose and lipid metabolism in mild hypercholesterolaemic individuals.

    PubMed

    Giacco, R; Clemente, G; Luongo, D; Lasorella, G; Fiume, I; Brouns, F; Bornet, F; Patti, L; Cipriano, P; Rivellese, A A; Riccardi, G

    2004-06-01

    The intake of 10 g/day of short-chain-fructo-oligosaccharides (sc-FOS) has been shown to increase significantly bifidus counts and to produce high amounts of short-chain fatty acids (SCFA), presumed to influence glucose and lipid metabolism. To evaluate the effects of moderate intake of sc-FOS on glucose and lipid metabolism in individuals with mild hypercholesterolaemia. A randomized double-blind sequential cross-over study. Thirty subjects of both genders (20 M/10 F), mean age 45.5+/-9.9 years (M+/-SD), BMI 26.6+/-2.2 kg/m(2), with plasma cholesterol >5.17 and <7.76 mmol/l and plasma triglycerides <3.45 mmol/l, participated in the study. The study was performed after a wash-out period of 1 month and a run-in period of 1 month to stabilize patients on a standard diet (CHO 50%, fat 30%, protein 20%, fibre 20 g/day) plus placebo (maltodextrine plus aspartame 15 g/day). At the end of run-in, subjects were randomly assigned to receive sc-FOS (Actilight) (10.6g/day) or placebo (maltodextrine plus aspartame 15 g/day) with tea and/or coffee for a duration of 2 months and thereafter switched to the other treatment for additional 2 months. Plasma glucose, total and lipoprotein (VLDL, LDL, HDL) cholesterol and triglyceride concentrations were measured in the fasting state at the end of run-in and of each treatment period. At the end of the two treatment periods, patients consumed a standard test meal (protein 15%, carbohydrate 34%, fat 51%, kJ 3988) 1h after the administration of 5.3g of sc-FOS or placebo; plasma glucose, insulin, free fatty acid (FFA) and triglyceride responses to the test meal were evaluated. No significant difference in fasting parameters was detected between the two treatments. After sc-FOS and placebo plasma cholesterol levels were, respectively, 6.47+/-0.70 and 6.44+/-0.78 mmol/l (n.s.) and plasma triglycerides were 1.53+/-0.71 and 1.56+/-0.53 mmol/l (n.s.). No significant differences were observed in cholesterol and triglyceride content of VLDL, LDL

  5. Plasma lipids and fatty acid synthase activity are regulated by short-chain fructo-oligosaccharides in sucrose-fed insulin-resistant rats.

    PubMed

    Agheli, N; Kabir, M; Berni-Canani, S; Petitjean, E; Boussairi, A; Luo, J; Bornet, F; Slama, G; Rizkalla, S W

    1998-08-01

    The aim of this study was to evaluate the chronic effects of a short-chain fructo-oligosaccharide (FOS)-containing diet on plasma lipids and the activity of fatty acid synthase (FAS) in insulin-resistant rats. Normal male Sprague-Dawley rats, 5 wk old, were randomly assigned to two groups and fed either a sucrose-rich diet (S, 575 g sucrose /kg diet and 140 g lipids/kg diet) or a sucrose-rich diet supplemented with 10 g/100 g short-chain fructo-oligosaccharides (S/FOS). A third reference group (R) was fed a standard nonpurified diet (g/kg, 575 g starch, 50 g fat). After 3 wk the sucrose-fed rats (compared with the R group) were characterized by the following: 1) higher insulin responses after a glucose challenge (P < 0.05); 2) heavier liver (P < 0.001) and retroperitoneal adipose tissue (P < 0.01); 3) hypertriglyceridemia (P < 0.0001) and higher plasma free fatty acids (P < 0.0001); and 4) higher fatty acid synthase activity in the liver but a low activity in the adipose tissue (P < 0.001). The addition of FOS to the diet resulted in 11% lower liver weight than in the S group (P < 0.05) and tended to result in lower adipose tissue weight (P < 0.11). Plasma triglycerides and plasma free fatty acids were lower in S/FOS- than in S-fed rats (P < 0.05). Chylomicrons + VLDL, and intermediate density lipoprotein (IDL) concentrations did not differ between groups, nor was plasma cholesterol influenced by diet. Hepatic FAS activity was lower in S/FOS-fed rats than in the S-fed rats (P < 0.05). In adipose tissue, however, this activity tended to be greater in rats fed S/FOS than in rats fed the S diet (P < 0.07). In conclusion, in a rat model of diet-induced (57.5% sucrose and 14% lipids) insulin resistance, the addition of short-chain FOS prevented some lipid disorders, lowered fatty acid synthase activity in the liver and tended to raise this activity in the adipose tissue. Short-chain FOS, in addition to being a nondigestible sweetener with good bulking capacity, might be

  6. Molecular assessment of the fecal microbiota in healthy cats and dogs before and during supplementation with fructo-oligosaccharides (FOS) and inulin using high-throughput 454-pyrosequencing

    PubMed Central

    Barcenas-Walls, Jose R.; Suchodolski, Jan S.; Steiner, Jörg M.

    2017-01-01

    Prebiotics are selectively fermentable dietary compounds that result in changes in the composition and/or activity of the intestinal microbiota, thus conferring benefits upon host health. In veterinary medicine, commercially available products containing prebiotics have not been well studied with regard to the changes they trigger on the composition of the gut microbiota. This study evaluated the effect of a commercially available nutraceutical containing fructo-oligosaccharides (FOS) and inulin on the fecal microbiota of healthy cats and dogs when administered for 16 days. Fecal samples were collected at two time points before and at two time points during prebiotic administration. Total genomic DNA was obtained from fecal samples and 454-pyrosequencing was used for 16S rRNA gene bacterial profiling. The linear discriminant analysis (LDA) effect size (LEfSe) method was used for detecting bacterial taxa that may respond (i.e., increase or decrease in its relative abundance) to prebiotic administration. Prebiotic administration was associated with a good acceptance and no side effects (e.g., diarrhea) were reported by the owners. A low dose of prebiotics (50 mL total regardless of body weight with the end product containing 0.45% of prebiotics) revealed a lower abundance of Gammaproteobacteria and a higher abundance of Veillonellaceae during prebiotic administration in cats, while Staphylococcaceae showed a higher abundance during prebiotic administration in dogs. These differences were not sufficient to separate bacterial communities as shown by analysis of weighted UniFrac distance metrics. A predictive approach of the fecal bacterial metagenome using Phylogenetic Investigation of Communities by Reconstruction of Unobserved States (PICRUSt) also did not reveal differences between the period before and during prebiotic administration. A second trial using a higher dose of prebiotics (3.2 mL/kg body weight with the end product containing 3.1% of prebiotics) was

  7. A short-term ingestion of fructo-oligosaccharides increases immunoglobulin A and mucin concentrations in the rat cecum, but the effects are attenuated with the prolonged ingestion.

    PubMed

    Komura, Mika; Fukuta, Tomonori; Genda, Tomomi; Hino, Shingo; Aoe, Seiichiro; Kawagishi, Hirokazu; Morita, Tatsuya

    2014-01-01

    We examined the effects of fructo-oligosaccharides (FOS) on IgA and mucin secretion in the rat cecum after different ingestion periods. Rats were fed a control diet or a diet containing FOS for 1, 2, 4, and 8 wk. FOS ingestion greatly increased IgA and mucin concentrations at 1 and 2 wk, but the effects were disappeared or attenuated at 4 and 8 wk. After 1 wk, FOS induced higher lactobacilli and lactate concentrations and lower cecal pH in the cecum, but the alterations were moderated with the prolonged ingestion accompanying with increasing short-chain fatty acid concentrations. At 1 and 2 wk, FOS increased IgA plasma cells and polymeric immunoglobulin receptor expression in the cecal mucosa and strongly depressed fecal mucinase activities related to the lower cecal pH. These findings may explain the FOS-induced early elevation of IgA and mucin. Clearly, FOS effects on IgA and mucin secretion considerably differ depending on the ingestion period.

  8. Fatty Acid Composition and Sensory Characteristics of Eggs Obtained from Hens Fed Flaxseed Oil, Dried Whitebait and/or Fructo-oligosaccharide

    PubMed Central

    Yi, Haechang; Hwang, Keum Taek; Regenstein, Joe M.; Shin, Sung Woo

    2014-01-01

    This study was conducted to assess the effects of flaxseed oil and dried whitebait as a source of ω-3 fatty acids (ω-3 FA), which could be used to produce eggs enriched with ω-3 FA, and of fructo-oligosaccharide (FOS) as a source of prebiotics on performance of hens (commercial Hy-Line Brown laying hens), and FA composition, internal quality, and sensory characteristics of the eggs. Dietary FOS increased egg weight. The amounts of α-linolenic (ALA), eicosapentaenoic (EPA) and docosahexaenoic acids (DHA) in the eggs from the hens fed the flaxseed oil alone or flaxseed oil+dried whitebait diets were higher than those of the control. Hedonic scores for off-flavor, fishy flavor, buttery taste and overall acceptability of the eggs from the hens fed the diet containing flaxseed oil+ dried whitebait were lower (p<0.05) than those of the control. Overall acceptability of the eggs from the hens fed the diet containing soybean oil+dried whitebait was lower (p<0.05) than that of the control. However, all the sensory attributes of the eggs from the hens fed the diet containing flaxseed oil, dried whitebait and FOS were not significantly different from those of the control. These results confirmed that flaxseed oil increases the ALA content in the eggs and a combination of flaxseed oil and dried whitebait increases EPA and DHA in the eggs. Of significance was that addition of FOS to the flaxseed oil+dried whitebait diet improves the sensory characteristics of the eggs enriched with ω-3 FA. PMID:25050046

  9. Appetite during consumption of enteral formula as a sole source of nutrition: the effect of supplementing pea-fibre and fructo-oligosaccharides.

    PubMed

    Whelan, Kevin; Efthymiou, Loukia; Judd, Patricia A; Preedy, Victor R; Taylor, Moira A

    2006-08-01

    Liquid enteral formulas are commonly used as a sole source of nutritional support of patients in hospital and community settings. Their effect on appetite has important consequences for dietary management of such patients and is likely to be affected by the formula composition. The aim of the present study was to compare appetite within healthy subjects consuming both a standard formula and one supplemented with pea-fibre (10 g/l) and fructo-oligosaccharide (FOS; 5 g/l) as a sole source of nutrition. Eleven healthy subjects consumed a standard formula or a pea-fibre/FOS formula as a sole source of nutrition for 14 d in a double-blind, cross-over trial. Appetite was recorded using standard 100 mm lines anchored at each end by a phrase denoting the most extreme appetite sensation. Consumption of the pea-fibre/FOS formula resulted in higher mean fullness (46 v. 37 mm, P=0.035), minimum fullness (13 v. 9 mm, P=0.024) and minimum satiety (12 v. 8 mm, P=0.012) compared to the standard formula. As there were no differences in macronutrient intake between formulas, these differences are likely to be due to supplementation with pea-fibre and FOS. The effect on appetite of the composition of an enteral formula, both with respect to nutrient content and functional components such as pea-fibre and FOS, may be an important aspect to consider in the dietary management of patients consuming enteral formula as a sole source of nutrition.

  10. Digestive tolerance and postprandial glycaemic and insulinaemic responses after consumption of dairy desserts containing maltitol and fructo-oligosaccharides in adults

    PubMed Central

    Respondek, F; Hilpipre, C; Chauveau, P; Cazaubiel, M; Gendre, D; Maudet, C; Wagner, A

    2014-01-01

    Background/objectives: To evaluate the short-term digestive tolerance and glycaemic response of several associations of maltitol and short-chain fructo-oligosaccharides (scFOS) used to replace sugars (for example, dextrose) in foods. Subjects/methods: Thirty-six healthy subjects aged 18–60 years were recruited for the study and 32 completed it. The subjects consumed six different mixtures of dextrose, maltitol and scFOS added in a chocolate dairy dessert at a dosage of 35 g. The test days were separated by 2-week washout periods. The subjects reported the intensity of four individual gastrointestinal (GI) symptoms, number of bowel movements and stool frequency for the 48 h following consumption of the dessert. A subgroup of 18 subjects also provided blood samples 2 h after intake to evaluate the postprandial glycaemic and insulinaemic responses. Results: The composite score calculated from the intensity of flatulence, borborygmi, bloating and discomfort was significantly higher (P<0.0001) for all the desserts containing maltitol and/or scFOS than for the control dessert containing dextrose, but remains at the level of mild effects. The number of bowel movements was also slightly increased (P=0.0006) and the stools were softer (P=0.0045) for the first 24 h but not after (P=0.1373 and 0.5420, respectively). Blood glycaemic and insulinaemic responses were lower for all the sugar-free recipes containing maltitol and scFOS in comparison to the control one (P<0.0001). Conclusions: This study has shown that maltitol and scFOS can be used jointly when formulating sugar-free foods with the benefit to lower postprandial glycaemic response with only a small and transient increase in non-serious GI symptoms. PMID:24642779

  11. Fructo-oligosaccharides and iron bioavailability in anaemic rats: the effects on iron species distribution, ferroportin-1 expression, crypt bifurcation and crypt cell proliferation in the caecum.

    PubMed

    Lobo, Alexandre R; Gaievski, Eduardo H S; De Carli, Eduardo; Alvares, Eliana P; Colli, Célia

    2014-10-28

    The present study investigated the effects of fructo-oligosaccharides (FOS) on the bioavailability of Fe from ferric pyrophosphate (FP), a water-insoluble compound, in Fe-deficient anaemic rats that were subjected to a Hb repletion assay. Male Wistar rats (n 64) were fed adequate or low (8 mg/kg) Fe diets for 15 d followed by 1 or 2 weeks of Fe repletion with diets providing 35 mg Fe/kg as ferrous sulphate (FS), FP or FP that was mixed with 7·5% FOS in the form of yacon flour or Raftilose P95 (RAF), a purified source of FOS. The effects of FOS were observed within the 1st week of the repletion period. Fe bioavailability was improved by FOS supplementation, as measured by Hb regeneration efficiency and hepatic Fe stores, which were more pronounced in the RAF group. Moreover, RAF supplementation resulted in a higher biological value relative to that of the FP group. FOS supplementation resulted in caecal enlargement, in addition to acidification and Fe species redistribution in the caecal contents relative to the control rats. These effects occurred concomitantly with decreased ferroportin (FPN)-1 expression in the caecal mucosa, which was similar in magnitude to that observed in the FS group. Caecum mucosal morphometry was influenced by FOS supplementation, whereas crypt fission and cell proliferation were highest in the caecum of the RAF group. These results reinforce the effects of FOS as Fe bioavailability enhancers in anaemic rats that are sustained by early changes in their caecal environment (decreased mucosal FPN-1 expression and increased Fe absorbability, crypt fission and cellularity).

  12. Fatty Acid Composition and Sensory Characteristics of Eggs Obtained from Hens Fed Flaxseed Oil, Dried Whitebait and/or Fructo-oligosaccharide.

    PubMed

    Yi, Haechang; Hwang, Keum Taek; Regenstein, Joe M; Shin, Sung Woo

    2014-07-01

    This study was conducted to assess the effects of flaxseed oil and dried whitebait as a source of ω-3 fatty acids (ω-3 FA), which could be used to produce eggs enriched with ω-3 FA, and of fructo-oligosaccharide (FOS) as a source of prebiotics on performance of hens (commercial Hy-Line Brown laying hens), and FA composition, internal quality, and sensory characteristics of the eggs. Dietary FOS increased egg weight. The amounts of α-linolenic (ALA), eicosapentaenoic (EPA) and docosahexaenoic acids (DHA) in the eggs from the hens fed the flaxseed oil alone or flaxseed oil+dried whitebait diets were higher than those of the control. Hedonic scores for off-flavor, fishy flavor, buttery taste and overall acceptability of the eggs from the hens fed the diet containing flaxseed oil+ dried whitebait were lower (p<0.05) than those of the control. Overall acceptability of the eggs from the hens fed the diet containing soybean oil+dried whitebait was lower (p<0.05) than that of the control. However, all the sensory attributes of the eggs from the hens fed the diet containing flaxseed oil, dried whitebait and FOS were not significantly different from those of the control. These results confirmed that flaxseed oil increases the ALA content in the eggs and a combination of flaxseed oil and dried whitebait increases EPA and DHA in the eggs. Of significance was that addition of FOS to the flaxseed oil+dried whitebait diet improves the sensory characteristics of the eggs enriched with ω-3 FA.

  13. Effects of supplementation with a calcium-rich marine-derived multi-mineral supplement and short-chain fructo-oligosaccharides on serum lipids in postmenopausal women.

    PubMed

    Cronin, Barbara E; Allsopp, Philip J; Slevin, Mary M; Magee, Pamela J; Livingstone, M Barbara E; Strain, J J; McSorley, Emeir M

    2016-02-28

    Recent literature suggests that Ca supplements have adverse effects on cardiovascular health. The effects of a Ca-rich supplement administered alone or in combination with short-chain fructo-oligosaccharides (scFOS) on serum lipids in postmenopausal women were examined using secondary data from a 24-month double-blind randomised controlled study. A total of 300 postmenopausal women were randomly assigned to daily supplements of 800 mg of Ca (2·4 g Aquamin) (Ca), 800 mg of Ca with 3 g of scFOS (CaFOS) or control (maltodextrin) (MD). A full lipid profile, body composition, blood pressure and a range of cytokines were measured at baseline and after 24 months. Intention-to-treat ANCOVA assessed treatment effects between the groups. A significant time-by-treatment effect was observed for LDL and total cholesterol for the Ca and CaFOS groups, with both groups having lower LDL and total cholesterol concentrations compared with MD after 24 months. The control group had mean (5·2 mmol/l) total cholesterol concentrations above the normal range (≤ 5 mmol/l) at 24 months, whereas values remained within the normal range in the treatment groups. There was no significant treatment effect on HDL-cholesterol, TAG, body composition, blood pressure or cytokine concentrations at 24 months, with the exception of IL-4, where there was a significant increase in the CaFOS group compared with the placebo. This study demonstrates a lipid-lowering effect of both the Ca-rich supplement alone and the supplement with scFOS. At the 4-year follow-up, there was no significant difference between the groups for reported diagnosed cardiovascular conditions.

  14. Normal Growth of Healthy Infants Born from HIV+ Mothers Fed a Reduced Protein Infant Formula Containing the Prebiotics Galacto-Oligosaccharides and Fructo-Oligosaccharides: A Randomized Controlled Trial

    PubMed Central

    da Costa Ribeiro, Hugo; Ribeiro, Tereza Cristina Medrado; de Mattos, Angela Peixoto; Pontes, Mariana; Sarni, Roseli Oselka Saccardo; Cruz, Maria Letícia Santos; Nogueira-de-Almeida, Carlos Alberto; Mussi-Pinhata, Marisa M; de Carvalho Norton, Rocksane; Steenhout, Philippe

    2015-01-01

    OBJECTIVE The aim of the current study was to evaluate the safety of a new reduced protein (2.1 g/100 kcal) infant formula containing 4 g/L of 90% galacto-oligosaccharides (GOS) and 10% fructo-oligosaccharides (FOS). METHODS Healthy term infants from Brazil were enrolled. Those born to human immunodeficiency virus (HIV)-positive mothers were randomized to a test (n = 65) or control (n = 63) formula group. Infants born to HIV-negative mothers were either exclusively breast-fed (n = 79) or received a mixed diet (breast milk and test formula, n = 65). Between 2 weeks and 4 months of age, infants were exclusively fed according to their assigned group. Anthropometric measurements were taken at baseline, 1, 2, 3, 4, 6, 8, 10, and 12 months. Digestive tolerance was evaluated during the first 4 months. The primary outcome was mean daily weight gain between 2 weeks and 4 months in the test formula and breast-fed groups. RESULTS Data from all infants (N = 272) were used in the intention-to-treat (ITT) analysis and data from 230 infants were used in the per-protocol (PP) analysis. The difference in mean daily weight gain between 2 weeks and 4 months in the test formula and breast-fed groups was 1.257 g/day (one-sided 95% confidence interval [CI]: −0.705 to inf, P < 0.001) in the PP analysis, showing that the lower bound of the 95% CI was above the −3.0 g/day non-inferiority margin. Results were similar in the ITT analysis. Symptoms of digestive tolerance and frequency of adverse events were similar in the two groups. CONCLUSIONS The formula containing 2.1 g/100 kcal protein and GOS and FOS was safe and tolerated well. PMID:25788839

  15. Short-chain fructo-oligosaccharide and inulin modulate inflammatory responses and microbial communities in Caco2-bbe cells and in a mouse model of intestinal injury.

    PubMed

    Johnson-Henry, Kathene C; Pinnell, Lee J; Waskow, Alexandra M; Irrazabal, Thergiory; Martin, Alberto; Hausner, Martina; Sherman, Philip M

    2014-11-01

    Few studies have focused on the ability of prebiotics to prevent pathogen-induced cellular changes or alter the composition of the intestinal microbiota in complimentary relevant cell and animal models of inflammatory bowel disease. The objective of this study was to determine if pretreatment with inulin and a short-chain fructo-oligosaccharide (sc-FOS) prevents enterohemorrhagic Escherichia coli (EHEC) O157:H7 infection in Caco2-bbe epithelial cells and what effect 10% wt:v sc-FOS or inulin has on C57BL/6 mice under sham conditions or pretreatment with prebiotics before Citrobacter rodentium infection (10(8) colony-forming units). Actin rearrangement and tight junction protein (zona occludin-1) were examined with immunofluorescence. Barrier function was assessed by a fluorescent probe and by measuring transepithelial electrical resistance (TER). Alterations in cytokine gene expression and microbiome were assessed with quantitative reverse transcriptase-polymerase chain reaction and fluorescence in situ hybridization. Short-chain fatty acids (SCFAs) were measured by GC. sc-FOS added to monolayers altered actin polymerization without affecting TER or permeability to a fluorescein isothiocyanate (FITC) probe, whereas inulin increased TER (P < 0.005) and altered actin arrangement without affecting FITC permeability. Neither prebiotic attenuated EHEC-induced decreases in barrier function. Prebiotics increased interleukin 10 (Il10) and transforming growth factor-β (Tgfβ) cytokine responses alone (P < 0.05) or with EHEC O157:H7 infection (P < 0.05) in vitro. Increases in tumor necrosis factor-α (Tnfα) (P < 0.05) and decreases in chemokine CXC motif ligand 8 (Cxcl8) (P < 0.05) expression were observed with prebiotic treatment prior to EHEC infection. No differences were noted in barrier function or cytokine responses in the absence or presence of C. rodentium in vivo. Alterations in microbiome were evident at 6 d and 10 d postinfection in treatment groups, but a

  16. In vitro determination of prebiotic properties of oligosaccharides derived from an orange juice manufacturing by-product stream.

    PubMed

    Manderson, K; Pinart, M; Tuohy, K M; Grace, W E; Hotchkiss, A T; Widmer, W; Yadhav, M P; Gibson, G R; Rastall, R A

    2005-12-01

    Fermentation properties of oligosaccharides derived from orange peel pectin were assessed in mixed fecal bacterial culture. The orange peel oligosaccharide fraction contained glucose in addition to rhamnogalacturonan and xylogalacturonan pectic oligosaccharides. Twenty-four-hour, temperature- and pH-controlled, stirred anaerobic fecal batch cultures were used to determine the effects that oligosaccharides derived from orange products had on the composition of the fecal microbiota. The effects were measured through fluorescent in situ hybridization to determine changes in bacterial populations, fermentation end products were analyzed by high-performance liquid chromatography to assess short-chain fatty acid concentrations, and subsequently, a prebiotic index (PI) was determined. Pectic oligosaccharides (POS) were able to increase the bifidobacterial and Eubacterium rectale numbers, albeit resulting in a lower prebiotic index than that from fructo-oligosaccharide metabolism. Orange albedo maintained the growth of most bacterial populations and gave a PI similar to that of soluble starch. Fermentation of POS resulted in an increase in the Eubacterium rectale numbers and concomitantly increased butyrate production. In conclusion, this study has shown that POS can have a beneficial effect on the fecal microflora; however, a classical prebiotic effect was not found. An increase in the Eubacterium rectale population was found, and butyrate levels increased, which is of potential benefit to the host.

  17. A Specific Mixture of Fructo-Oligosaccharides and Bifidobacterium breve M-16V Facilitates Partial Non-Responsiveness to Whey Protein in Mice Orally Exposed to β-Lactoglobulin-Derived Peptides

    PubMed Central

    Kostadinova, Atanaska I.; Meulenbroek, Laura A. P. M.; van Esch, Betty C. A. M.; Hofman, Gerard A.; Garssen, Johan; Willemsen, Linette E. M.; Knippels, Léon M. J.

    2017-01-01

    Oral tolerance is a promising approach for allergy prevention in early life, but it strongly depends on allergen exposure and proper immune environment. Small tolerance-inducing peptides and dietary immunomodulatory components may comprise an attractive method for allergy prevention in at-risk infants. This study aimed to investigate whether early oral exposure to β-lactoglobulin-derived peptides (BLG-peptides) and a specific synbiotic mixture of short- and long- chain fructo-oligosaccharides (scFOS/lcFOS, FF) and Bifidobacterium breve (Bb) M-16V (FF/Bb) can prevent cow’s milk allergy (CMA). Three-week-old female C3H/HeOuJ mice were orally exposed to phosphate buffered saline (PBS), whey protein, or a mixture of four synthetic BLG-peptides combined with a FF/Bb-enriched diet prior to intragastric sensitization with whey protein and cholera toxin. To assess the acute allergic skin response and clinical signs of allergy, mice were challenged intradermally with whole whey protein. Serum immunoglobulins were analyzed after a whey protein oral challenge. Cytokine production by allergen-reactivated splenocytes was measured and changes in T cells subsets in the spleen, mesenteric lymph nodes, and intestinal lamina propria were investigated. Pre-exposing mice to a low dosage of BLG-peptides and a FF/Bb-enriched diet prior to whey protein sensitization resulted in a significant reduction of the acute allergic skin response to whey compared to PBS-pretreated mice fed a control diet. Serum immunoglobulins were not affected, but anaphylactic symptom scores remained low and splenocytes were non-responsive in whey-induced cytokine production. In addition, preservation of the Th1/Th2 balance in the small intestine lamina propria was a hallmark of the mechanism underlying the protective effect of the BLG-peptides–FF/Bb intervention. Prior exposure to BLG-peptides and a FF/Bb-enriched diet is a promising approach for protecting the intestinal Th1/Th2 balance and reducing the

  18. Enzymatic production of specifically distributed hyaluronan oligosaccharides.

    PubMed

    Yuan, Panhong; Lv, Mengxian; Jin, Peng; Wang, Miao; Du, Guocheng; Chen, Jian; Kang, Zhen

    2015-09-20

    High-molecular-mass hyaluronan (HA) was controllably depolymerized in pure aqueous solution with recombinant leech hyaluronidase (HAase). The HAase concentration per unit HA and hydrolysis time played important roles in molecular mass distribution. By modulating the concentrations of HAase and controlling the hydrolysis time, any molar-mass-defined HA oligomers could be efficiently and specifically produced on a large scale (40 g/L), such as HA oligosaccharides with weight-average molar mass of 4000, 10,000, and 30,000Da and end hydrolysates containing only HA6 and HA4. High performance liquid chromatography-size exclusion chromatography, polyacrylamide gel electrophoresis, capillary zone electrophoresis, and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry confirmed low polydispersity of the produced molar-mass-defined HA oligosaccharides. Therefore, large-scale production of defined HA oligosaccharides with narrow molecular mass distribution will significantly promote progress in related research and its potential applications. Copyright © 2015 Elsevier Ltd. All rights reserved.

  19. Oligosaccharides: state of the art.

    PubMed

    Delzenne, N M

    2003-02-01

    Oligosaccharides, consisting of a mixture of hexose oligomers with a variable extent of polymerisation, are food products with interesting nutritional properties. They may be naturally present in food, mostly in fruits, vegetables or grains, or produced by biosynthesis from natural sugars or polysaccharides and added to food products because of their nutritional properties or organoleptic characteristics. The dietary intake of oligosaccharides is difficult to estimate, but it may reach 3-13 g/d per person (for fructo-oligosaccharides), depending on the population. The extent of resistance to enzymic reactions occurring in the upper part of the gastrointestinal tract allows oligosaccharides to become 'colonic nutrients', as some intestinal bacterial species express specific hydrolases and are able to convert oligosaccharides into short-chain fatty acids (acetate, lactate, propionate, butyrate) and/or gases by fermentation. Oligosaccharides that selectively promote some interesting bacterial species (e.g. lactobacilli, bifidobacteria), and thus equilibrate intestinal microflora, are now termed prebiotics. The pattern of short-chain fatty acid production in the caeco-colon, as well as the prebiotic effect, if demonstrated, are dynamic processes that vary with the type of oligosaccharide (e.g. extent of polymerisation, nature of hexose moieties), the duration of the treatment, the initial composition of flora or the diet in which they are incorporated. Experimental data obtained in vitro and in vivo in animals, and also recent data obtained in human subjects, support the involvement of dietary oligosaccharides in physiological processes in the different intestinal cell types (e.g. mucins production, cell division, immune cells function, ionic transport) and also outside the gastrointestinal tract (e.g. hormone production, lipid and carbohydrates metabolism). The present paper gives an overview of the future development of oligosaccharides, newly recognised as dietary

  20. Profiles of human milk oligosaccharides and production of some human milk oligosaccharides in transgenic animals.

    PubMed

    Prieto, Pedro Antonio

    2012-05-01

    During the decade of the 1990s and the first years of the current century, our group embarked on a project to study and synthesize human milk oligosaccharides. This report describes 2 unexpected collateral observations from that endeavor. The first observation was the detection and confirmation of 2 rare neutral human milk oligosaccharides profiles that were uncovered while assessing oligosaccharide content in hundreds of samples of human milk. One of these lacked fucosylated structures altogether, and the other lacked the oligosaccharide 3-fucosyllactose [Galβ1-4(Fucα1-3)Glc]. We used glycoconjugate probes to determine whether the unusual profiles were mirrored by fucosylation of milk glycoproteins. The results show that the lack of fucosylated oligosaccharides in these samples corresponds to the absence of equivalent fucosylated motifs in milk glycoproteins. The second finding was a shortened and distinct lactation process in transgenic rabbits expressing the human fucosyltransferase 1. During the first day of lactation, these animals expressed milk that contained both lactose and 2'-fucosylactose, but on the second day, the production of milk was severely diminished, and by the fourth day, no lactose was detected in their milk. Meanwhile, the concentration of fucosylated glycoproteins increased from the onset of lactation through its premature termination. These 2 findings may shed light on the glycobiology of milk and perhaps on mammary gland differentiation.

  1. Profiles of Human Milk Oligosaccharides and Production of Some Human Milk Oligosaccharides in Transgenic Animals12

    PubMed Central

    Prieto, Pedro Antonio

    2012-01-01

    During the decade of the 1990s and the first years of the current century, our group embarked on a project to study and synthesize human milk oligosaccharides. This report describes 2 unexpected collateral observations from that endeavor. The first observation was the detection and confirmation of 2 rare neutral human milk oligosaccharides profiles that were uncovered while assessing oligosaccharide content in hundreds of samples of human milk. One of these lacked fucosylated structures altogether, and the other lacked the oligosaccharide 3-fucosyllactose [Galβ1–4(Fucα1–3)Glc]. We used glycoconjugate probes to determine whether the unusual profiles were mirrored by fucosylation of milk glycoproteins. The results show that the lack of fucosylated oligosaccharides in these samples corresponds to the absence of equivalent fucosylated motifs in milk glycoproteins. The second finding was a shortened and distinct lactation process in transgenic rabbits expressing the human fucosyltransferase 1. During the first day of lactation, these animals expressed milk that contained both lactose and 2′-fucosylactose, but on the second day, the production of milk was severely diminished, and by the fourth day, no lactose was detected in their milk. Meanwhile, the concentration of fucosylated glycoproteins increased from the onset of lactation through its premature termination. These 2 findings may shed light on the glycobiology of milk and perhaps on mammary gland differentiation. PMID:22585925

  2. Production of oligosaccharides in yogurt containing bifidobacteria and yogurt cultures.

    PubMed

    Lamoureux, L; Roy, D; Gauthier, S F

    2002-05-01

    Yogurts were prepared by using yogurt cultures combined to mixed cultures of bifidobacteria (Bifidobacterium animalis, Bifidobacterium bifidum, Bifidobacterium breve, Bifidobacterium infantis, and Bifidobacterium longum) and by adding a preincubation step (1.5 h at 50 degrees C) with bifidobacteria to the conventional method of manufacture in order to produce oligosaccharides. The survival of bifidobacteria was drastically affected during storage of yogurts, except for products containing B. animalis, in which viable counts remained at >10(6) cfu/g after 28 d of storage at 4 degrees C. Oligosaccharides with a degree of polymerization of 3 were produced during the preincubation step (0.31 to 0.68%), and the amount in the final products varied according to the species of bifidobacteria inoculated during the preincubation step or the concentration of bifidobacteria used as second inoculum during the fermentation process. In fact, the higher concentration of oligosaccharides measured at the end of the fermentation process (0.72%) and the 28 d-storage period (0.67%) was obtained for yogurts containing B. infantis. However, yogurts containing B. breve showed higher beta-galactosidase activities and had lower lactose concentrations after the fermentation process and the storage period than the other yogurts. The use of a mixed cultures of bifidobacteria (B. animalis, B. infantis, or B. breve) thus allows the production of yogurts in which bifidobacteria can survive in relatively high cell numbers and contain appreciable amount of oligosaccharides.

  3. Galactoglucomannan Oligosaccharide Supplementation Affects Nutrient Digestibility, Fermentation End-Product Production, and Large Bowel Microbiota of the Dog

    USDA-ARS?s Scientific Manuscript database

    A galactoglucomannan oligosaccharide (GGMO) obtained from fiberboard production was evaluated as a dietary supplement for dogs. The GGMO substrate contained high concentrations of oligosaccharides containing mannose, xylose, and glucose, with the mannose component accounting for 35% of dry matter. ...

  4. Production of arabinoxylan-oligosaccharides from flaxseed (Linum usitatissimum).

    PubMed

    Guilloux, Karine; Gaillard, Isabelle; Courtois, Josiane; Courtois, Bernard; Petit, Emmanuel

    2009-12-09

    Flaxseed mucilage from Linum usitatissimum L. species was constituted by arabinoxylan (about 75%) and pectin (about 25%). A new procedure was developed to obtain only arabinoxylans which implicated treatment of the pectin fraction by enzymatic hydrolysis with pectinase. Then three processes of depolymerization were evaluated on arabinoxylans. First, a thermic hydrolysis in mild acid conditions was performed and an ultrafiltration process was used as purification method. Second, the potential of xylanases from different glycoside hydrolase families for arabinoxylan-oligosaccharides (AXOS) production was tested, and finally a radical depolymerization was conducted. Average molecular weights were determined by high pressure size exclusion chromatography coupled with multiple angle laser light scattering (MALLS), and carbohydrate compositions were determined by high pH anion exchange chromatography pulse amperometric detector (HPAEC-PAD). Both chemical and enzymatic treatments were inefficient to convert arabinoxylans from flaxseed mucilage into AXOS. Only radical depolymerization process was allowed to obtain arabinoxylan-oligosaccharides presenting different molecular weights (11.9 x 10(3) to 1.9 x 10(3) g mol(-1)) with satisfactory yields (75% to 35%).

  5. Enzymatic production of pectic oligosaccharides from onion skins.

    PubMed

    Babbar, Neha; Baldassarre, Stefania; Maesen, Miranda; Prandi, Barbara; Dejonghe, Winnie; Sforza, Stefano; Elst, Kathy

    2016-08-01

    Onion skins are evaluated as a new raw material for the enzymatic production of pectic oligosaccharides (POS) with a targeted degree of polymerization (DP). The process is based on a two-stage process consisting of a chelator-based crude pectin extraction followed by a controlled enzymatic hydrolysis. Treatment of the extracted crude onion skin's pectin with various enzymes (EPG-M2, Viscozyme and Pectinase) shows that EPG-M2 is the most appropriate enzyme for tailored POS production. The experiments reveal that the highest amount of DP2 and DP3 is obtained at a time scale of 75-90min with an EPG-M2 concentration of 26IU/mL. At these conditions the production amounts 2.5-3.0% (w/w) d.m for DP2 and 5.5-5.6% (w/w) d.m for DP3 respectively. In contrast, maximum DP4 production of 5.2-5.5% (w/w) d.m. is obtained with 5.2IU/mL at a time scale of 15-30min. Detailed LC-MS analysis reveals the presence of more methylated oligomers compared to acetylated forms in the digests. Copyright © 2016 Elsevier Ltd. All rights reserved.

  6. Functional oligosaccharides: application and manufacture.

    PubMed

    Rastall, R A

    2010-01-01

    Oligosaccharides are attracting increasing interest as prebiotic functional food ingredients. They can be extracted or obtained by enzymatic hydrolysis from a variety of biomass sources or synthesized from simple oligosaccharides by enzymatic transfer reactions. The major prebiotic oligosaccharides on the market are inulin, fructo-oligosaccharides, and galacto-oligosaccharides. They have been evaluated using a range of in vitro and in vivo methods, although there is a need for more large-scale human trials using modern microbiological methods. Prebiotics are being studied for their effects on gut health and well being and specific clinical conditions, including colon cancer, inflammatory bowel disease (IBD), acute infections, and mineral absorption. Developing understanding of the functional ecology of the human gut is influencing current thinking on what a prebiotic might achieve and is providing new targets for prebiotic intervention.

  7. Galactoglucomannan oligosaccharide Supplementation affects Nutrient Digestibility, Fermentation End-product Production, and Large Bowel Microbiota of the Dog

    USDA-ARS?s Scientific Manuscript database

    A galactoglucomannan oligosaccharide (GGMO) obtained from fiberboard production was evaluated as a dietary supplement for dogs. The GGMO substrate contained high concentrations of mannose, xylose, and glucose oligosaccharides. Adult dogs assigned to a 6x6 Latin square design were fed six diets, ea...

  8. Pectic oligosaccharides from agricultural by-products: production, characterization and health benefits.

    PubMed

    Babbar, Neha; Dejonghe, Winnie; Gatti, Monica; Sforza, Stefano; Elst, Kathy

    2016-08-01

    Pectin containing agricultural by-products are potential sources of a new class of prebiotics known as pectic oligosaccharides (POS). In general, pectin is made up of homogalacturonan (HG, α-1,4-linked galacturonic acid monomers) and rhamnogalacturonan (RG, alternate galacturonic acid and rhamnose backbone with neutral side chains). Controlled hydrolysis of pectin containing agricultural by-products like sugar beet, apple, olive and citrus by chemical, enzymatic and hydrothermal can be used to produce oligo-galacturonides (GalpOS), galacto-oligosaccharides (GalOS), rhamnogalacturonan-oligosaccharides (RGOS), etc. However, extensive research is needed to establish the role of POS, both as a prebiotic as well as therapeutic agent. This review comprehensively covers different facets of POS, including the nature and chemistry of pectin and POS, potential agricultural residual sources of pectin, pre-treatment methods for facilitating selective extraction of pectin, identification and characterization of POS, health benefits and important applications of POS in food and feed. This review has been compiled to establish a platform for future research in the purification and characterization of POS and for in vivo and in vitro studies of important POS, so that they could be commercially exploited.

  9. Feruloylated and nonferuloylated arabino-oligosaccharides from sugar beet pectin selectively stimulate the growth of Bifidobacterium spp. in human fecal in vitro fermentations.

    PubMed

    Holck, Jesper; Lorentzen, Andrea; Vigsnæs, Louise K; Licht, Tine R; Mikkelsen, Jørn D; Meyer, Anne S

    2011-06-22

    The side chains of the rhamnogalacturonan I fraction in sugar beet pectin are particularly rich in arabinan moieties, which may be substituted with feruloyl groups. In this work the arabinan-rich fraction resulting from sugar beet pulp based pectin production was separated by Amberlite XAD hydrophobic interaction and membrane separation into four fractions based on feruloyl substitution and arabino-oligosaccharide chain length: short-chain (DP 2-10) and long-chain (DP 7-14) feruloylated and nonferuloylated arabino-oligosaccharides, respectively. HPAEC, SEC, and MALDI-TOF/TOF analyses of the fractions confirmed the presence of singly and doubly substituted feruloylated arabino-oligosaccharides in the feruloyl-substituted fractions. In vitro microbial fermentation by human fecal samples (n = 6 healthy human volunteers) showed a selective stimulation of bifidobacteria by both the feruloylated and the nonferuloylated long-chain arabino-oligosaccharides to the same extent as the prebiotic fructo-oligosaccharides control. None of the fractions stimulated the growth of the potential pathogen Clostridium difficile in monocultures. This work provides a first report on the separation of potentially bioactive feruloylated arabino-oligosaccharides from sugar beet pulp and an initial indication of the potentially larger bifidogenic effect of relatively long-chain arabino-oligosaccharides as opposed to short-chain arabino-oligosaccharides.

  10. Effect of cranberry (Vaccinium macrocarpon) oligosaccharides on the formation of advanced glycation end-products

    PubMed Central

    Sun, Jiadong; Liu, Weixi; Ma, Hang; Marais, Jannie P. J.; Khoo, Christina; Dain, Joel A.; Rowley, David C.; Seeram, Navindra P.

    2016-01-01

    BACKGROUND: The formation and accumulation of advanced glycation end-products (AGEs) are implicated in several chronic human illnesses including type-2 diabetes, renal failure, and neurodegenerative diseases. The cranberry (Vaccinium macrocarpon) fruit has been previously reported to show anti-AGEs effects, attributed primarily to its phenolic constituents. However, there is lack of similar data on the non-phenolic constituents found in the cranberry fruit, in particular, its carbohydrate constituents. Herein, a chemically characterized oligosaccharide-enriched fraction purified from the cranberry fruit was evaluated for its potential anti-AGEs and free radical scavenging effects. OBJECTIVE: The aim of this study was to evaluate the in vitro anti-AGEs and free radical scavenging effects of a chemically characterized oligosaccharide-enriched fraction purified from the North American cranberry (Vaccinium macrocarpon) fruit. METHOD: The cranberry oligosaccharide-enriched fraction was purified from cranberry hull powder and characterized based on spectroscopic and spectrometric (NMR, MALDI-TOF-MS, and HPAEC-PAD) data. The oligosaccharide-enriched fraction was evaluated for its anti-AGEs and free radical scavenging effects by the bovine serum albumin-fructose, and DPPH assays, respectively. RESULTS: Fractionation of cranberry hull material yielded an oligosaccharide-enriched fraction named Cranf1b-CL. The 1H NMR and MALDI-TOF-MS revealed that Cranf1b-CL consists of oligosaccharides ranging primarily from 6-mers to 9-mers. The monosaccharide composition of Cranf1b-CL was arabinose (25%), galactose (5%), glucose (47%) and xylose (23%). In the bovine serum albumin-fructose assay, Cranf1b-CL inhibited AGEs formation in a concentration-dependent manner with comparable activity to the synthetic antiglycating agent, aminoguanidine, used as the positive control (57 vs. 75%; both at 500μg/mL). In the DPPH free radical scavenging assay, Cranf1b-CL showed superior activity to the

  11. Effect of cranberry (Vaccinium macrocarpon) oligosaccharides on the formation of advanced glycation end-products.

    PubMed

    Sun, Jiadong; Liu, Weixi; Ma, Hang; Marais, Jannie P J; Khoo, Christina; Dain, Joel A; Rowley, David C; Seeram, Navindra P

    2016-06-16

    BACKGROUND: The formation and accumulation of advanced glycation end-products (AGEs) are implicated in several chronic human illnesses including type-2 diabetes, renal failure, and neurodegenerative diseases. The cranberry (Vaccinium macrocarpon) fruit has been previously reported to show anti-AGEs effects, attributed primarily to its phenolic constituents. However, there is lack of similar data on the non-phenolic constituents found in the cranberry fruit, in particular, its carbohydrate constituents. Herein, a chemically characterized oligosaccharide-enriched fraction purified from the cranberry fruit was evaluated for its potential anti-AGEs and free radical scavenging effects. OBJECTIVE: The aim of this study was to evaluate the in vitro anti-AGEs and free radical scavenging effects of a chemically characterized oligosaccharide-enriched fraction purified from the North American cranberry (Vaccinium macrocarpon) fruit. METHOD: The cranberry oligosaccharide-enriched fraction was purified from cranberry hull powder and characterized based on spectroscopic and spectrometric (NMR, MALDI-TOF-MS, and HPAEC-PAD) data. The oligosaccharide-enriched fraction was evaluated for its anti-AGEs and free radical scavenging effects by the bovine serum albumin-fructose, and DPPH assays, respectively. RESULTS: Fractionation of cranberry hull material yielded an oligosaccharide-enriched fraction named Cranf1b-CL. The (1)H NMR and MALDI-TOF-MS revealed that Cranf1b-CL consists of oligosaccharides ranging primarily from 6-mers to 9-mers. The monosaccharide composition of Cranf1b-CL was arabinose (25%), galactose (5%), glucose (47%) and xylose (23%). In the bovine serum albumin-fructose assay, Cranf1b-CL inhibited AGEs formation in a concentration-dependent manner with comparable activity to the synthetic antiglycating agent, aminoguanidine, used as the positive control (57 vs. 75%; both at 500μg/mL). In the DPPH free radical scavenging assay, Cranf1b-CL showed superior activity to

  12. Evolved beta-galactosidases from Geobacillus stearothermophilus with improved transgalactosylation yield for galacto-oligosaccharide production.

    PubMed

    Placier, Gaël; Watzlawick, Hildegard; Rabiller, Claude; Mattes, Ralf

    2009-10-01

    A mutagenesis approach was applied to the beta-galactosidase BgaB from Geobacillus stearothermophilus KVE39 in order to improve its enzymatic transglycosylation of lactose into oligosaccharides. A simple screening strategy, which was based on the reduction of the hydrolysis of a potential transglycosylation product (lactosucrose), provided mutant enzymes possessing improved synthetic properties for the autocondensation product from nitrophenyl-galactoside and galacto-oligosaccharides (GOS) from lactose. The effects of the mutations on enzyme activity and kinetics were determined. An change of one arginine to lysine (R109K) increased the oligosaccharide yield compared to that for the wild-type BgaB. Subsequently, saturation mutagenesis at this position demonstrated that valine and tryptophan further increased the transglycosylation performance of BgaB. During the transglycosylation reaction with lactose of the evolved beta-galactosidases, a major trisaccharide was formed. Its structure was characterized as beta-D-galactopyranosyl-(1-->3)-beta-D-galactopyranosyl-(1-->4)-D-glucopyranoside (3'-galactosyl-lactose). At the lactose concentration of 18% (wt/vol), this trisaccharide was obtained in yields of 11.5% (wt/wt) with GP21 (BgaB R109K), 21% with GP637.2 (BgaB R109V), and only 2% with the wild-type BgaB enzyme. GP643.3 (BgaB R109W) was shown to be the most efficient mutant, with a 3'-galactosyl-lactose production of 23%.

  13. Assessing the effects of different prebiotic dietary oligosaccharides in sheep milk ice cream.

    PubMed

    Balthazar, C F; Silva, H L A; Vieira, A H; Neto, R P C; Cappato, L P; Coimbra, P T; Moraes, J; Andrade, M M; Calado, V M A; Granato, D; Freitas, M Q; Tavares, M I B; Raices, R S L; Silva, M C; Cruz, A G

    2017-01-01

    The objective of this study was to assess the effects of different prebiotic dietary oligosaccharides (inulin, fructo-oligosaccharide, galacto-oligossacaride, short-chain fructo-oligosaccharide, resistant starch, corn dietary oligosaccharide and polydextrose) in non-fat sheep milk ice cream processing through physical parameters, water mobility and thermal analysis. Overall, the fat replacement by dietary prebiotic oligosaccharides significantly decreased the melting time, melting temperature and the fraction and relaxation time for fat and bound water (T22) while increased the white intensity and glass transition temperature. The replacement of sheep milk fat by prebiotics in sheep milk ice cream constitutes an interesting option to enhance nutritional aspects and develop a functional food.

  14. Supercritical water treatment for cello-oligosaccharide production from microcrystalline cellulose.

    PubMed

    Tolonen, Lasse K; Juvonen, Minna; Niemelä, Klaus; Mikkelson, Atte; Tenkanen, Maija; Sixta, Herbert

    2015-01-12

    Microcrystalline cellulose was treated in supercritical water at 380 °C and at a pressure of 250 bar for 0.2, 0.4, and 0.6s. The yield of the ambient-water-insoluble precipitate and its average molar mass decreased with an extended treatment time. The highest yield of 42 wt% for DP2-9 cello-oligosaccharides was achieved after the 0.4s treatment. The reaction products included also 11 wt% ambient-water-insoluble precipitate with a DP(w) of 16, and 6.1 wt% monomeric sugars, and 37 wt% unidentified degradation products. Oligo- and monosaccharide-derived dehydration and retro-aldol fragmentation products were analyzed via a combination of HPAEC-PAD-MS, ESI-MS/MS, and GC-MS techniques. The total amount of degradation products increased with treatment time, and fragmented (glucosyl(n)-erythrose, glucosyl(n)-glycolaldehyde), and dehydrated (glucosyl(n)-levoglucosan) were identified as the main oligomeric degradation products from the cello-oligosaccharides.

  15. Optimization of galacto-oligosaccharide production by Bifidobacterium infantis RW-8120 using response surface methodology.

    PubMed

    Roy, D; Daoudi, L; Azaola, A

    2002-11-01

    Oligosaccharide (OS) production, cell concentration (2 x 10(9) colony-forming unit/ml), lactose concentration (25% wt/vol), reaction time (6 h), and temperature (50 degrees C) were chosen as the central condition of the central composite design (CCD) for optimizing the production process using Bifidobacterium infantis RW-8120 in skim milk. Statistical analysis (P<0.01) revealed that the most relevant variable concerning OS production and yield was the lactose concentration. The coefficient of determination (R(2)) is good for the second-order OS production model (0.92) and fairly good for the second-order nonlinear OS yield model (0.816). An increase of lactose concentration and temperature resulted in a higher OS production. The optimal values for OS production appear to be near the area associated with the central points of the modeling design except for the lactose concentration, which was 40% (wt/vol) of the final volume.

  16. Recyclable Strategy for the Production of High-Purity Galacto-oligosaccharides by Kluyveromyces lactis.

    PubMed

    Sun, Huaisheng; You, Shengping; Wang, Mengfan; Qi, Wei; Su, Rongxin; He, Zhimin

    2016-07-20

    A recyclable strategy for the production of high-purity (>95%) galacto-oligosaccharides (GOS) was developed using Kluyveromyces lactis in both the synthesis and purification steps. For the synthesis of GOS, ethanol-permeabilized cells (p-cells) of K. lactis were used because the enhanced permeability facilitated the mass transfer of the substrate and the release of oligosaccharide products. For the purification of GOS, non-permeabilized K. lactis cells (np-cells) were preferred as a result of their intrinsic cell membrane barrier toward GOS, which led to the selective consumption of carbohydrate. In this way, undesired glucose, galactose, and lactose in the raw GOS solution can be completely removed. This strategy is recyclable not only because of the high stability and reusability of p-cells and np-cells but also because the ethanol, which is simultaneously generated during the purification, can be reused for the preparation of p-cells. The strategy proposed in this study is a promising candidate for the efficient production of high-purity GOS.

  17. Enzyme catalysed production of sialylated human milk oligosaccharides and galactooligosaccharides by Trypanosoma cruzi trans-sialidase.

    PubMed

    Holck, Jesper; Larsen, Dorte M; Michalak, Malwina; Li, Haiying; Kjærulff, Louise; Kirpekar, Finn; Gotfredsen, Charlotte H; Forssten, Sofia; Ouwehand, Arthur C; Mikkelsen, Jørn D; Meyer, Anne S

    2014-03-25

    A Trypanosoma cruzi trans-sialidase (E.C. 3.2.1.18) was cloned into Pichia pastoris and expressed. The pH and temperature optimum of the enzyme was determined as pH 5.7 and 30°C. Using casein glycomacropeptide (CGMP) and lactose as sialyl-donor and acceptor respectively, the optimal donor/acceptor ratio for the trans-sialidase catalysed 3'-sialyllactose production was found to be 1:4. Quantitative amounts of 3'-sialyllactose were produced from CGMP and lactose at a yield of 40mg/g CGMP. The 3'-sialyllactose obtained exerted a stimulatory effect on selected probiotic strains, including different Bifidobacterium strains in single culture fermentations. The trans-sialidase also catalysed the transfer of sialic acid from CGMP to galacto-oligosaccharides (GOS) and to the human milk oligosaccharide (HMO) backbone lacto-N-tetraose (LNT) to produce 3'-sialyl-GOS, including doubly sialylated GOS products, and 3'-sialyl-LNT, respectively. This work thus provides proof of the concept of producing 3'-sialyllactose and potentially other sialylated HMOs as well as sialylated GOS enzymatically by trans-sialidase activity, while at the same time providing valorisation of CGMP, a co-processing product from cheese manufacture.

  18. Direct enzymatic production of oligosaccharide mixtures from sugar beet pulp: experimental evaluation and mathematical modeling.

    PubMed

    Martínez, Martina; Gullón, Beatriz; Yáñez, Remedios; Alonso, José Luis; Parajó, Juan Carlos

    2009-06-24

    The potential of sugar beet pulp as raw material for neutral and acidic oligosaccharide production by direct enzymatic treatment was evaluated. The effect of the polygalacturonase to solid ratio (PGaseSR), cellulase activity to polygalacturonase activity ratio (CPGaseR), and reaction time (t) on several dependent variables (selected to quantify the mass of recovered liquors, the conversion of each polysaccharide into monosaccharides, and the conversion of each polysaccharide into oligomers) was studied. Mathematical models suitable for reproducing and predicting the experimental results were also developed. Operational conditions leading to a maximum oligomer production were calculated from models being PGaseSR = 10 U/g, CPGaseR = 0.725 filter paper units/U, and t = 12.82 h. Under these conditions, the models predicted that 906 kg of liquors containing 26.7 kg of oligosaccharides can be obtained from 100 kg of SBP, the distribution being as follows: 5.9 kg of glucooligosaccharides, 2.4 kg of galactooligosaccharides, 9.7 kg of arabinooligosaccharides, and 8.7 kg of oligogalacturonides. Therefore, this study demonstrated that pectic oligomers (for which prebiotic properties have been reported) can be obtained from SBP at high yield by direct enzymatic hydrolysis using mixtures of cellulases and pectinases.

  19. Galacto-oligosaccharides and Colorectal Cancer: Feeding our Intestinal Probiome

    PubMed Central

    Bruno-Barcena, Jose M.; Azcarate-Peril, M. Andrea

    2014-01-01

    Prebiotics are ingredients selectively fermented by the intestinal microbiota that promote changes in the microbial community structure and/or their metabolism, conferring health benefits to the host. Studies show that β (1–4) galacto-oligosaccharides [β (1–4) GOS], lactulose and fructo-oligosaccharides increase intestinal concentration of lactate and short chain fatty acids, and stool frequency and weight, and they decrease fecal concentration of secondary bile acids, fecal pH, and nitroreductase and β-glucuronidase activities suggesting a clear role in colorectal cancer (CRC) prevention. This review summarizes research on prebiotics bioassimilation, specifically β (1–4) GOS, and their potential role in CRC. We also evaluate research that show that the impact of prebiotics on host physiology can be direct or through modulation of the gut intestinal microbiome, specifically the probiome (autochtonous beneficial bacteria), we present studies on a potential role in CRC progression to finally describe the current state of β (1–4) GOS generation for industrial production. PMID:25584074

  20. Comparative structural characterization of 7 commercial galacto-oligosaccharide (GOS) products.

    PubMed

    van Leeuwen, Sander S; Kuipers, Bas J H; Dijkhuizen, Lubbert; Kamerling, Johannis P

    2016-04-29

    Many β-galactosidase enzymes convert lactose into a mixture of galacto-oligosaccharides (GOS) when incubated under the right conditions. Recently, the composition of commercial Vivinal GOS produced by Bacillus circulans β-galactosidase was studied in much detail in another study by van Leeuwen et al. As a spin-off of this study, we used the developed analytical strategy for the evaluation of 6 anonymous commercial GOS products, in comparison with Vivinal GOS. These GOS products were first subjected to HPLC-SEC, calibrated HPAEC-PAD profiling (glucose units in relation to a malto-oligosaccharide ladder), and 1D (1)H NMR spectroscopy. For a more detailed analysis and support of the conclusions based on the initial analysis, the GOS products were separated into DP-pure subpools on Bio-Gel P-2 (MALDI-TOF-MS analysis), which were subjected to calibrated HPAEC-PAD profiling and (1)H NMR analysis. Unidentified peaks from different GOS products, not present in Vivinal GOS, were isolated for detailed structural characterization. In this way, the differences between the various GOS products in terms of DP distribution and type of glycosidic linkages were established. A total of 13 new GOS structures were characterized, adding structural-reporter-group signals and HPAEC-PAD based glucose unit G.U. values to the analytical toolbox. The newly characterized products enhance the quality of the database with GOS structures up to DP4. The combined data provide a firm basis for the rapid profiling of the GOS products of microbial β-galactosidase enzymes. Copyright © 2016 Elsevier Ltd. All rights reserved.

  1. Novel α-L-Fucosidases from a Soil Metagenome for Production of Fucosylated Human Milk Oligosaccharides.

    PubMed

    Lezyk, Mateusz; Jers, Carsten; Kjaerulff, Louise; Gotfredsen, Charlotte H; Mikkelsen, Maria D; Mikkelsen, Jørn D

    2016-01-01

    This paper describes the discovery of novel α-L-fucosidases and evaluation of their potential to catalyse the transglycosylation reaction leading to production of fucosylated human milk oligosaccharides. Seven novel α-L-fucosidase-encoding genes were identified by functional screening of a soil-derived metagenome library and expressed in E. coli as recombinant 6xHis-tagged proteins. All seven fucosidases belong to glycosyl hydrolase family 29 (GH 29). Six of the seven α-L-fucosidases were substrate-inhibited, moderately thermostable and most hydrolytically active in the pH range 6-7, when tested with para-nitrophenyl-α-L-fucopyranoside (pNP-Fuc) as the substrate. In contrast, one fucosidase (Mfuc6) exhibited a high pH optimum and an unusual sigmoidal kinetics towards pNP-Fuc substrate. When tested for trans-fucosylation activity using pNP-Fuc as donor, most of the enzymes were able to transfer fucose to pNP-Fuc (self-condensation) or to lactose. With the α-L-fucosidase from Thermotoga maritima and the metagenome-derived Mfuc5, different fucosyllactose variants including the principal fucosylated HMO 2'-fucosyllactose were synthesised in yields of up to ~6.4%. Mfuc5 was able to release fucose from xyloglucan and could also use it as a fucosyl-donor for synthesis of fucosyllactose. This is the first study describing the use of glycosyl hydrolases for the synthesis of genuine fucosylated human milk oligosaccharides.

  2. Comparison of chito-oligosaccharide production from three different colloidal chitosans using the endochitonsanolytic system of Bacillus thuringiensis.

    PubMed

    Olicón-Hernández, Dario Rafael; Vázquez-Landaverde, Pedro Alberto; Cruz-Camarillo, Ramón; Rojas-Avelizapa, Luz Irene

    2017-02-07

    Bacillus thuringiensis is a nonhuman pathogen bacterium that is used as a fungal and insect biocontrol agent. Because of its environmental interaction, it possesses several extracellular enzymes that are able to degrade chitin and chitosan, two of the most important polymers because of their application in numerous fields. However, in recent years, it has been observed that oligosaccharides from the enzymatic degradation of chitosan have important benefits for human health. Comparison and exploration of the production of chito-oligosaccharides from different sources of chitosan will improve the process parameters and expand the biotechnology based in these molecules. This study shows the production of chito-oligosaccharides from three different sources of colloidal chitosan and conducts a qualitative-quantitative comparison between them, using the extracellular enzyme of B. thuringiensis. We found that in the three substrates, it is possible to get a mixture of chito-oligosaccharides from dimer to hexamer in a concentration range from 0.72 to 8.09 mg · g(-1) of original substrate. The best substrate to obtain these molecules was commercial chitosan as it has the highest production yields.

  3. Efficient free fatty acid production in engineered Escherichia coli strains using soybean oligosaccharides as feedstock.

    PubMed

    Wang, Dan; Wu, Hui; Thakker, Chandresh; Beyersdorf, Jared; Bennett, George N; San, Ka-Yiu

    2015-01-01

    To be competitive with current petrochemicals, microbial synthesis of free fatty acids can be made to rely on a variety of renewable resources rather than on food carbon sources, which increase its attraction for governments and companies. Industrial waste soybean meal is an inexpensive feedstock, which contains soluble sugars such as stachyose, raffinose, sucrose, glucose, galactose, and fructose. Free fatty acids were produced in this report by introducing an acyl-ACP carrier protein thioesterase and (3R)-hydroxyacyl-ACP dehydratase into E. coli. Plasmid pRU600 bearing genes involved in raffinose and sucrose metabolism was also transformed into engineered E. coli strains, which allowed more efficient utilization of these two kinds of specific oligosaccharide present in the soybean meal extract. Strain ML103 (pRU600, pXZ18Z) produced ~1.60 and 2.66 g/L of free fatty acids on sucrose and raffinose, respectively. A higher level of 2.92 g/L fatty acids was obtained on sugar mixture. The fatty acid production using hydrolysate obtained from acid or enzyme based hydrolysis was evaluated. Engineered strains just produced ~0.21 g/L of free fatty acids with soybean meal acid hydrolysate. However, a fatty acid production of 2.61 g/L with a high yield of 0.19 g/g total sugar was observed on an enzymatic hydrolysate. The results suggest that complex mixtures of oligosaccharides derived from soybean meal can serve as viable feedstock to produce free fatty acids. Enzymatic hydrolysis acts as a much more efficient treatment than acid hydrolysis to facilitate the transformation of industrial waste from soybean processing to high value added chemicals.

  4. Oligosaccharide sequencing based on exo- and endoglycosidase digestion and liquid chromatographic analysis of the products.

    PubMed

    Prime, S; Dearnley, J; Ventom, A M; Parekh, R B; Edge, C J

    1996-01-12

    Exo- and endoglycosidases are used to sequence oligosaccharides and give valuable information on the monosaccharide sequence, together with the anomericity, the stereochemistry, and in some cases, the substitution pattern of the monosaccharides. Both sequential and parallel methods of oligosaccharide sequencing are discussed.

  5. Novel α-L-Fucosidases from a Soil Metagenome for Production of Fucosylated Human Milk Oligosaccharides

    PubMed Central

    Lezyk, Mateusz; Jers, Carsten; Kjaerulff, Louise; Gotfredsen, Charlotte H.; Mikkelsen, Maria D.; Mikkelsen, Jørn D.

    2016-01-01

    This paper describes the discovery of novel α-L-fucosidases and evaluation of their potential to catalyse the transglycosylation reaction leading to production of fucosylated human milk oligosaccharides. Seven novel α-L-fucosidase-encoding genes were identified by functional screening of a soil-derived metagenome library and expressed in E. coli as recombinant 6xHis-tagged proteins. All seven fucosidases belong to glycosyl hydrolase family 29 (GH 29). Six of the seven α-L-fucosidases were substrate-inhibited, moderately thermostable and most hydrolytically active in the pH range 6–7, when tested with para-nitrophenyl-α-L-fucopyranoside (pNP-Fuc) as the substrate. In contrast, one fucosidase (Mfuc6) exhibited a high pH optimum and an unusual sigmoidal kinetics towards pNP-Fuc substrate. When tested for trans-fucosylation activity using pNP-Fuc as donor, most of the enzymes were able to transfer fucose to pNP-Fuc (self-condensation) or to lactose. With the α-L-fucosidase from Thermotoga maritima and the metagenome-derived Mfuc5, different fucosyllactose variants including the principal fucosylated HMO 2’-fucosyllactose were synthesised in yields of up to ~6.4%. Mfuc5 was able to release fucose from xyloglucan and could also use it as a fucosyl-donor for synthesis of fucosyllactose. This is the first study describing the use of glycosyl hydrolases for the synthesis of genuine fucosylated human milk oligosaccharides. PMID:26800369

  6. Purification, characterization, and prebiotic properties of pectic oligosaccharides from orange peel wastes.

    PubMed

    Gómez, Belén; Gullón, Beatriz; Remoroza, Connie; Schols, Henk A; Parajó, Juan C; Alonso, José L

    2014-10-08

    Pectic oligosaccharides (POS) were obtained by hydrothermal treatment of orange peel wastes (OPW) and purified by membrane filtration to yield a refined product containing 90 wt % of the target products. AraOS (DP 3-21), GalOS (DP 5-12), and OGalA (DP 2-12, with variable DM) were identified in POS mixtures, but long-chain products were also present. The prebiotic potential of the concentrate was assessed by in vitro fermentation using human fecal inocula. For comparative purposes, similar experiments were performed using orange pectin and commercial fructo-oligosaccharides (FOS) as substrates for fermentation. The dynamics of selected microbial populations was assessed by fluorescent in situ hybridization (FISH). Gas generation, pH, and short-chain fatty acid (SCFA) production were also measured. Under the tested conditions, all of the considered substrates were utilized by the microbiota, and fermentation resulted in increased numbers of all the bacterial groups, but the final profile of the microbial population depended on the considered carbon source. POS boosted particularly the numbers of bifidobacteria and lactobacilli, so that the ratio between the joint counts of both genera and the total cell number increased from 17% in the inocula to 27% upon fermentation. SCFA generation from POS fermentation was similar to that observed with FOS, but pectin fermentation resulted in reduced butyrate generation.

  7. Acceptor products of alternansucrase with gentiobiose. Production of novel oligosaccharides for food and feed and elimination of bitterness.

    PubMed

    Côté, Gregory L

    2009-01-26

    In the presence of suitable acceptor molecules, dextransucrase makes a homologous series of oligosaccharides in which the isomers differ by a single glucosyl unit, whereas alternansucrase synthesizes one trisaccharide, two tetrasaccharides, etc. Previously, we showed that alternansucrase only forms certain isomers of DP>4 from maltose in measurable amounts, and that these oligosaccharides belong to the oligoalternan series rather than the oligodextran series. We now demonstrate that the acceptor products from gentiobiose, also formed in good yields (nearly 90% in unoptimized reactions), follow a pattern similar to those formed from maltose. The initial product is a single trisaccharide, alpha-d-Glcp-(1-->6)-beta-d-Glcp-(1-->6)-d-Glc. Two tetrasaccharides were formed in approximately equal quantities: alpha-d-Glcp-(1-->3)-alpha-d-Glcp-(1-->6)-beta-d-Glcp-(1-->6)-d-Glc and alpha-d-Glcp-(1-->6)-alpha-d-Glcp-(1-->6)-beta-d-Glcp-(1-->6)-d-Glc. Just one pentasaccharide was isolated from the reaction mixture, alpha-d-Glcp-(1-->6)-alpha-d-Glcp-(1-->3)-alpha-d-Glcp-(1-->6)-beta-d-Glcp-(1-->6)-d-Glc. Our hypothesis that the enzyme is incapable of forming two consecutive alpha-(1-->3) linkages, and does not form products with more than two consecutive alpha-(1-->6) linkages, apparently applies to other acceptors as well as to maltose. The glucosylation of gentiobiose reduces or eliminates its bitter taste.

  8. Heat treatment of curdlan enhances the enzymatic production of biologically active β-(1,3)-glucan oligosaccharides.

    PubMed

    Kumagai, Yuya; Okuyama, Masayuki; Kimura, Atsuo

    2016-08-01

    Biologically active β-(1,3)-glucan oligosaccharides were prepared from curdlan using GH64 enzyme (KfGH64). KfGH64 showed low activity toward native curdlan; thereby pretreatment conditions of curdlan were evaluated. KfGH64 showed the highest activity toward curdlan with heat treatment. The most efficient pretreatment (90°C for 0.5h) converted approximately 60% of curdlan into soluble saccharides under the optimized enzyme reaction conditions (pH 5.5, 37°C, 100rpm mixing speed, 24h, and 10μg of KfGH64/1g of curdlan). The resulting products were predominantly laminaripentaose and a small amount of β-(1,3)-glucans with an average degree of polymerization (DP) of 13 and 130. The products did not contain small oligosaccharides (DP<5), indicating that the hydrolysis of heat-treated curdlan by KfGH64 is a suitable method for the production of biologically active β-(1,3)-glucan oligosaccharides. Copyright © 2016 Elsevier Ltd. All rights reserved.

  9. Production of lactulose oligosaccharides by isomerisation of transgalactosylated cheese whey permeate obtained by β-galactosidases from dairy Kluyveromyces.

    PubMed

    Padilla, Beatriz; Frau, Florencia; Ruiz-Matute, Ana Isabel; Montilla, Antonia; Belloch, Carmela; Manzanares, Paloma; Corzo, Nieves

    2015-08-01

    β-Galactosidases from Kluyveromyces lactis and Kluyveromyces marxianus isolated from artisanal ewes' milk cheeses, were used to transgalactosylate lactose from cheese whey permeate (WP). The content of galactooligosaccharides (GOS) obtained by transgalactosylation was comparable with that formed using pure lactose as substrate. In order to obtain a mixture with higher prebiotic oligosaccharide content, isomerisation of the transgalactosylated WP was carried out using sodium aluminate as catalyst. The transgalactosylated mixtures at 6 h of reaction contained amounts of prebiotic carbohydrates (tagatose, lactulose, GOS and oligosaccharides derived from lactulose, OsLu) close to 50 g/100 g of total carbohydrates for all the strains tested, corresponding to 322 g prebiotics/kg whey permeate. Thus, the suitability of this methodology to produce mixtures of dietary non-digestible carbohydrates with prebiotic properties from WP has been demonstrated, which is interesting for the food industry since it increases the value and the applicability of this by-product from cheese manufacture.

  10. Rapid Screening of Bovine Milk Oligosaccharides in a Whey Permeate Product and Domestic Animal Milks by Accurate Mass Database and Tandem Mass Spectral Library.

    PubMed

    Lee, Hyeyoung; Cuthbertson, Daniel J; Otter, Don E; Barile, Daniela

    2016-08-17

    A bovine milk oligosaccharide (BMO) library, prepared from cow colostrum, with 34 structures was generated and used to rapidly screen oligosaccharides in domestic animal milks and a whey permeate powder. The novel library was entered into a custom Personal Compound Database and Library (PCDL) and included accurate mass, retention time, and tandem mass spectra. Oligosaccharides in minute-sized samples were separated using nanoliquid chromatography (nanoLC) coupled to a high resolution and sensitive quadrupole-Time of Flight (Q-ToF) MS system. Using the PCDL, 18 oligosaccharides were found in a BMO-enriched product obtained from whey permeate processing. The usefulness of the analytical system and BMO library was further validated using milks from domestic sheep and buffaloes. Through BMO PCDL searching, 15 and 13 oligosaccharides in the BMO library were assigned in sheep and buffalo milks, respectively, thus demonstrating significant overlap between oligosaccharides in bovine (cow and buffalo) and ovine (sheep) milks. This method was shown to be an efficient, reliable, and rapid tool to identify oligosaccharide structures using automated spectral matching.

  11. Enhancement of diosgenin production in Dioscorea zingiberensis cell cultures by oligosaccharides from its endophytic fungus Fusarium oxysporum Dzf17.

    PubMed

    Li, Peiqin; Mao, Ziling; Lou, Jingfeng; Li, Yan; Mou, Yan; Lu, Shiqiong; Peng, Youliang; Zhou, Ligang

    2011-12-19

    The effects of the oligosaccharides from the endophytic fungus Fusarium oxysporum Dzf17 as elicitors on diosgenin production in cell suspension cultures of its host Dioscorea zingiberensis were investigated. Three oligosaccharides, DP4, DP7 and DP10, were purified from the oligosaccharide fractions DP2-5, DP5-8 and DP8-12, respectively, which were prepared from the water-extracted mycelial polysaccharide of the endophytic fungus F. oxysporum Dzf17. When the cell cultures were treated with fraction DP5-8 at 20 mg/L on day 26 and harvested on day 32, the maximum diosgenin yield (2.187 mg/L) was achieved, which was 5.65-fold of control (0.387 mg/L). When oligosaccharides DP4, DP7 and DP10 were individually added to 26-day-old D. zingiberensis cell cultures at concentrations of 2, 4, 6, 8 and 10 mg/L in medium, DP7 at 6 mg/L was found to significantly enhance diosgenin production, with a yield of 3.202 mg/L, which was 8.27-fold of control. When the cell cultures were treated with DP7 twice on days 24 and 26, and harvested on day 30, both diosgenin content and yield were significantly increased and reached the maximums of 1.159 mg/g dw and 4.843 mg/L, both of which were higher than those of single elicitation, and were 9.19- and 12.38-fold of control, respectively.

  12. Selective chemical oxidation and depolymerization of switchgrass [corrected] (Panicum virgatum L.) xylan with [corrected] oligosaccharide product analysis by mass spectrometry.

    PubMed

    Bowman, Michael J; Dien, Bruce S; O'Bryan, Patricia J; Sarath, Gautam; Cotta, Michael A

    2011-04-15

    Xylan is a barrier to enzymatic hydrolysis of plant cell walls. It is well accepted that the xylan layer needs to be removed to efficiently hydrolyze cellulose; consequently, pretreatment conditions are (in part) optimized for maximal xylan depolymerization or displacement. Xylan consists of a long chain of β-1,4-linked xylose units substituted with arabinose (typically α-1,3-linked in grasses) and glucuronic acid (α-1,2-linked). Xylan has been proposed to have a structural function in plants and therefore may play a role in determining biomass reactivity to pretreatment. It has been proposed that substitutions along xylan chains are not random and, based upon studies of pericarp xylan, are organized in domains that have specific structural functions. Analysis of intact xylan is problematic because of its chain length (> degree of polymerization (d.p.) 100) and heterogeneous side groups. Traditionally, enzymatic end-point products have been characterized due to the limited products generated. Analysis of resultant arabino-xylo-oligosaccharides by mass spectrometry is complicated by the isobaric pentose sugars that primarily compose xylan. In this report, the variation in pentose ring structures was exploited for selective oxidation of the arabinofuranose primary alcohols followed by acid depolymerization to provide oligosaccharides with modified arabinose branches intact. Switchgrass samples were analyzed by hydrophilic interaction chromatography (HILIC)-liquid chromatography (LC)-mass spectrometry/mass spectrometry (MSMS) and off-line nanospray MS to demonstrate the utility of this chemistry for determination of primary hydroxyl groups on oligosaccharide structures, with potential applications for determining the sequence of arabino-xylo-oligosaccharides present in plant cell wall material.

  13. Bifidogenic effect and stimulation of short chain fatty acid production in human faecal slurry cultures by oligosaccharides derived from lactose and lactulose.

    PubMed

    Cardelle-Cobas, Alejandra; Fernández, María; Salazar, Nuria; Martínez-Villaluenga, Cristina; Villamiel, Mar; Ruas-Madiedo, Patricia; de los Reyes-Gavilán, Clara G

    2009-08-01

    Bifidogenic effect and stimulation of short chain fatty acid (SCFA) production by fractions of oligosaccharides with a DP> or =3 and Gal beta(1-6) linkages synthesised from lactose or lactulose by Pectinex Ultra SP-L and Lactozym 3000 L HP G were evaluated in human faecal slurries. Results were compared with those obtained for the commercial oligosaccharide mixture Vivinal-GOS. Quantitative real-time PCR showed that all galacto-oligosaccharide (GOS) fractions stimulated slightly higher bifidobacteria growth than lactose, lactulose and Vivinal-GOS. GOS fractions promoted the production of total SCFA and acetic acid in a similar way to Vivinal-GOS and more than glucose, lactose and lactulose. In conclusion, oligosaccharides derived from lactose and lactulose may represent a suitable alternative to lactulose for the design of new functional food ingredients.

  14. Galactoglucomannan oligosaccharide supplementation affects nutrient digestibility, fermentation end-product production, and large bowel microbiota of the dog.

    PubMed

    Faber, T A; Hopkins, A C; Middelbos, I S; Price, N P; Fahey, G C

    2011-01-01

    A galactoglucomannan oligosaccharide (GGMO) obtained from fiberboard production was evaluated as a dietary supplement for dogs. The GGMO substrate contained increased concentrations of oligosaccharides containing mannose, xylose, and glucose, with the mannose component accounting for 35% of DM. Adult dogs assigned to a 6 × 6 Latin square design were fed 6 diets, each containing a different concentration of supplemental GGMO (0, 0.5, 1, 2, 4, and 8%) that replaced dietary cellulose. Total tract DM and OM apparent digestibilities increased (P < 0.001) linearly, whereas total tract CP apparent digestibility decreased (P < 0.001) linearly as dietary GGMO substrate concentration increased. Fecal concentrations of acetate, propionate, and total short-chain fatty acids increased (P ≤ 0.001) linearly, whereas butyrate concentration decreased (P ≤ 0.001) linearly with increasing dietary concentrations of GGMO. Fecal pH decreased (P ≤ 0.001) linearly as dietary GGMO substrate concentration increased, whereas fecal score increased quadratically (P ≤ 0.001). Fecal phenol (P ≤ 0.05) and indole (P ≤ 0.01) concentrations decreased linearly with GGMO supplementation. Fecal biogenic amine concentrations were not different among treatments except for phenylethylamine, which decreased (P < 0.001) linearly as dietary GGMO substrate concentration increased. Fecal microbial concentrations of Escherichia coli, Lactobacillus spp., and Clostridium perfringens were not different among treatments. A quadratic increase (P ≤ 0.01) was noted for Bifidobacterium spp. as dietary GGMO substrate concentration increased. The data suggest positive nutritional properties of supplemental GGMO when incorporated in a good-quality dog food.

  15. Oligosaccharides in Food and Agriculture

    NASA Astrophysics Data System (ADS)

    Collins, Michelle E.; Rastall, Robert A.

    Oligosaccharides are an integral part of the daily diet for humans and animals. They are primarily used for their nutritional properties, however they are currently receiving much attention due to their physiological effect on the microflora of the gastrointestinal tract. Galacto-oligosaccharides and the fructan-type oligosaccharides, namely FOS and inulin are well established as beneficial to the host and are classified as prebiotic based on data from clinical studies. These compounds dominate this sector of the market, although there are oligosaccharides emerging which have produced very interesting in vitro results in terms of prebiotic status and human trials are required to strengthen the claim. Such compounds include pectic oligosaccharides, gluco-oligosaccharides, gentio-oligosaccharides, kojio-oligosaccharides, and alternan oligosaccharides. The raw materials for production of these prebiotic compounds are derived from natural sources such as plants but also from by products of the food processing industry. In addition to being prebiotic these compounds can be incorporated into foodstuffs due to the physiochemical properties they possess.

  16. Ethanol production from xylo-oligosaccharides by xylose-fermenting Saccharomyces cerevisiae expressing β-xylosidase.

    PubMed

    Fujii, Tatsuya; Yu, Guoce; Matsushika, Akinori; Kurita, Asami; Yano, Shinichi; Murakami, Katsuji; Sawayama, Shigeki

    2011-01-01

    Construction of xylose- and xylo-oligosaccharide-fermenting Saccharomyces cerevisiae strains is important, because hydrolysates derived from lignocellulosic biomass contain significant amounts of these sugars. We have obtained recombinant S. cerevisiae strain MA-D4 (D-XKXDHXR), expressing xylose reductase, xylitol dehydrogenase and xylulokinase. In the present study, we generated recombinant strain D-XSD/XKXDHXR by transforming MA-D4 with a β-xylosidase gene cloned from the filamentous fungus Trichoderma reesei. The intracellular β-xylosidase-specific activity of D-XSD/XKXDHXR was high, while that of the control strain was under the limit of detection. D-XSD/XKXDHXR produced ethanol, and xylose accumulated in the culture supernatant under fermentation in a medium containing xylo-oligosaccharides as sole carbon source. β-Xylosidase-specific activity in D-XSD/XKXDHXR declined due to xylose both in vivo and in vitro. D-XSD/XKXDHXR converted xylo-oligosaccharides in an enzymatic hydrolysate of eucalyptus to ethanol. These results indicate that D-XSD/XKXDHXR efficiently converted xylo-oligosaccharides to xylose and subsequently to ethanol.

  17. Oligosaccharides from land plants and algae: production and applications in therapeutics and biotechnology.

    PubMed

    Courtois, Josiane

    2009-06-01

    Since the past decades, oligosaccharides are considered for their potential biological activities. To exploit them, it was essential to obtain pure molecules in large amounts. Several strategies were developed to produce specific sugar sequences with specific substitution patterns from land plants and algae polysaccharides. Then, pure oligosaccharides were analyzed for their potential biological activities and relations between oligomers structure and function were tackled. First they can be health beneficial molecules when they are added to the diet to enhance the growth of probiotic bacteria, in that case, oligomers that resist to the digestive process are used as specific substrate for the growth of health beneficial bacteria. In other cases, oligomers have to interact with receptors on cells. In this instance, a specific conformation is needed to allow the sugar sequence to establish specific linkages with the receptor. So, to be adapted to the receptor, the oligosaccharides have to present specific groups to the receptor, there, the polymerization degree of oligosaccharides as well as the flexibility of the glycosidic linkages has to be considered.

  18. Production and bioactivity of pectic oligosaccharides from fruit and vegetable biomass

    USDA-ARS?s Scientific Manuscript database

    Pectin is abundant in various agro-industrial bio-resources such as citrus peel, apple pomace, cranberry pulp and sugar beet pulp. These materials can therefore be considered as a source of potential bioactive pectic oligosaccharides. This chapter reviews the various extraction and purification meth...

  19. Xylo-oligosaccharides production by autohydrolysis of corn fiber separated from DDGS

    USDA-ARS?s Scientific Manuscript database

    Xylo-oligosaccharides (XOS) are reported to have beneficial health properties, and are considered to be functional food ingredients. XOS was produced using corn fiber separated from distillers dried grains with solubles (DDGS). Corn fiber was treated with deionized water in a Parr-reactor, at temper...

  20. Production of Feruloyated Arabinoxylo-oligosaccharides from Maize (Zea mays) Bran by Microwave-Assisted Autohydrolysis

    USDA-ARS?s Scientific Manuscript database

    Maize bran was treated with microwave irradiation (160 – 200 °C for 2 – 20 min) to release feruolyated arabinoxylo-oligosaccharides (AXOS). Lower temperatures and shorter treatment times were consistent with low AXOS yields, while higher temperatures and longer reaction times also resulted in low y...

  1. In vitro digestion and fermentation properties of linear sugar-beet arabinan and its oligosaccharides.

    PubMed

    Moon, Jin Seok; Shin, So Yeon; Choi, Hye Sun; Joo, Wooha; Cho, Seung Kee; Li, Ling; Kang, Jung-Hyun; Kim, Tae-Jip; Han, Nam Soo

    2015-10-20

    This study was conducted to investigate the prebiotic effects of linear arabino-oligosaccharides (LAOS) and debranched (linear) sugar beet arabinan (LAR) for the development of new prebiotics. LAOS were prepared from LAR by enzymatic hydrolysis with endo-arabinanase from Bacillus licheniformis, followed by removal of the arabinose fraction by incubation with resting cells of Leuconostoc mesenteroides. The resulting LAOS contained DP2 (28.7%), DP3 (49.9%), DP4 (20.1%), and DP5 (1.16%). A standardized digestibility test showed that LAOS and LAR were not digestible. Individual cultures of 24 strains of gastrointestinal bacteria showed that LAOS and LAR stimulated growth of Lactobacillus brevis, Bifidobacterium longum, and Bacteroides fragilis. In vitro batch fermentation using human fecal samples showed that LAOS had higher bifidogenic properties than LAR; LAOS increased the population of bifidobacteria which produced short-chain fatty acids (SCFAs). LAOS was fermented slowly compared to fructo-oligosaccharides and this may permit SCFA production in the distal colon. This study demonstrates that LAOS prepared from LAR are promising dietary substrates for improvement of human intestinal health.

  2. Xylo-oligosaccharides enhance the growth of bifidobacteria and Bifidobacterium lactis in a simulated colon model.

    PubMed

    Mäkeläinen, H; Forssten, S; Saarinen, M; Stowell, J; Rautonen, N; Ouwehand, A C

    2010-03-01

    A semi-continuous, anaerobic colon simulator, with four vessels mimicking the conditions of the human large intestine, was used to study the fermentation of xylo-oligosaccharides (XOS). Three XOS compounds and a xylan preparation were fermented for 48 hours by human colonic microbes. Fructo-oligosaccharides (FOS) were used as a prebiotic reference. As a result of the fermentation, the numbers of Bifidobacterium increased in all XOS and xylan simulations when compared to the growth observed in the baseline simulations, and increased levels of Bifidobacterium lactis were measured with the two XOS compounds that had larger distribution of the degree of polymerisation. Fermentation of XOS and xylan increased the microbial production of short chain fatty acids in the simulator vessels; especially the amounts of butyrate and acetate were increased. XOS was more efficient than FOS in increasing the numbers of B. lactis in the colonic model, whereas FOS increased the Bifidobacterium longum numbers more. The selective fermentation of XOS by B. lactis has been demonstrated in pure culture studies, and these results further indicate that the combination of B. lactis and XOS would form a successful, selective synbiotic combination.

  3. Biotechnological production of fucosylated human milk oligosaccharides: Prokaryotic fucosyltransferases and their use in biocatalytic cascades or whole cell conversion systems.

    PubMed

    Petschacher, Barbara; Nidetzky, Bernd

    2016-10-10

    Human milk oligosaccharides (HMOs) constitute a class of complex carbohydrates unique to mother's milk and are strongly correlated to the health benefits of breastfeeding in infants. HMOs are important as functional ingredients of advanced infant formula and have attracted broad interest for use in health-related human nutrition. About 50% of the HMOs structures contain l-fucosyl residues, which are introduced into nascent oligosaccharides by enzymatic transfer from GDP-l-fucose. To overcome limitation in the current availability of fucosylated HMOs, biotechnological approaches for their production have been developed. Functional expression of the fucosyltransferase(s) and effective supply of GDP-l-fucose, respectively, are both bottlenecks of the biocatalytic routes of synthesis. Strategies of in vitro and in vivo production of fucosylated HMOs are reviewed here. Besides metabolic engineering for enhanced HMO production in whole cells, the focus is on the characteristics and the heterologous overexpression of prokaryotic α1,2- and α1,3/4-fucosyltransferases. Up to 20g/L of fucosylated HMOs were obtained in optimized production systems. Optimized expression enabled recovery of purified fucosyltransferases in a yield of up to 45mg/L culture for α1,2-fucosyltransferases and of up to 200mg protein/L culture for α1,3/4-fucosyltransferases.

  4. Monitoring and preparation of neoagaro- and agaro-oligosaccharide products by high performance anion exchange chromatography systems.

    PubMed

    Kazłowski, Bartosz; Pan, Chorng Liang; Ko, Yuan Tih

    2015-05-20

    A series of neoagaro-oligosaccharides (NAOS) were prepared by β-agarase digestion and agaro-oligosaccharides (AOS) by HCl hydrolysis from agarose with defined quantity and degree of polymerization (DP). Chain-length distribution in the crude product mixtures were monitored by two high performance anion exchange chromatography systems coupled with a pulsed amperometric detector. Method 1 utilized two separation columns: a CarboPac(™) PA1 and a CarboPac(™) PA100 connected in series and method 2 used the PA100 alone. Method 1 resolved the product in size ranges consisting of DP 1-46 for NAOS and DP 1-32 for AOS. Method 2 clearly resolved saccharide product sizes within DP 26. The optimized system utilizing a semi-preparative CarboPac(™) PA100 column was connected with a fraction collector to isolate and quantify individually separated products. This study established systems for the preparation and qualitative and quantitative measurements as well as for the isolation of various sizes of oligomers generated from agarose.

  5. Short-chain fatty acid formation in the hindgut of rats fed oligosaccharides varying in monomeric composition, degree of polymerisation and solubility.

    PubMed

    Nilsson, Ulf; Nyman, Margareta

    2005-11-01

    The contents of short-chain fatty acids were investigated in rats fed lactitol, lactulose and four fructo-oligosaccharides of different degree of polymerisation and solubility. Fructo-oligosaccharides with a low degree of polymerisation (2-8) generated the highest levels of butyric acid all along the hindgut, whereas fructo-oligosaccharides with a high degree of polymerisation (10-60) generated the highest levels of propionic acid. These specific differences were also generally reflected in the caecal pools and molar proportions of short-chain fatty acids. The lower solubility of the fructo-oligosaccharides was related to a lower degree of caecal fermentation. Lactulose and lactitol yielded high proportions of acetic acid and low proportions of butyric acid. It is concluded that both the degree of polymerisation and the solubility may affect short-chain fatty acid formation, whereas the fructose content per se seem to be of less importance. This may be of interest when designing foods with specific health effects.

  6. The characterisation of an alkali-stable maltogenic amylase from Bacillus lehensis G1 and improved malto-oligosaccharide production by hydrolysis suppression.

    PubMed

    Abdul Manas, Nor Hasmaliana; Pachelles, Samson; Mahadi, Nor Muhammad; Illias, Rosli Md

    2014-01-01

    A maltogenic amylase (MAG1) from alkaliphilic Bacillus lehensis G1 was cloned, expressed in Escherichia coli, purified and characterised for its hydrolysis and transglycosylation properties. The enzyme exhibited high stability at pH values from 7.0 to 10.0. The hydrolysis of β-cyclodextrin (β-CD) produced malto-oligosaccharides of various lengths. In addition to hydrolysis, MAG1 also demonstrated transglycosylation activity for the synthesis of longer malto-oligosaccharides. The thermodynamic equilibrium of the multiple reactions was shifted towards synthesis when the reaction conditions were optimised and the water activity was suppressed, which resulted in a yield of 38% transglycosylation products consisting of malto-oligosaccharides of various lengths. Thin layer chromatography and high-performance liquid chromatography analyses revealed the presence of malto-oligosaccharides with a higher degree of polymerisation than maltoheptaose, which has never been reported for other maltogenic amylases. The addition of organic solvents into the reaction further suppressed the water activity. The increase in the transglycosylation-to-hydrolysis ratio from 1.29 to 2.15 and the increased specificity toward maltopentaose production demonstrated the enhanced synthetic property of the enzyme. The high transglycosylation activity of maltogenic amylase offers a great advantage for synthesising malto-oligosaccharides and rare carbohydrates.

  7. The Characterisation of an Alkali-Stable Maltogenic Amylase from Bacillus lehensis G1 and Improved Malto-Oligosaccharide Production by Hydrolysis Suppression

    PubMed Central

    Abdul Manas, Nor Hasmaliana; Pachelles, Samson; Mahadi, Nor Muhammad; Illias, Rosli Md.

    2014-01-01

    A maltogenic amylase (MAG1) from alkaliphilic Bacillus lehensis G1 was cloned, expressed in Escherichia coli, purified and characterised for its hydrolysis and transglycosylation properties. The enzyme exhibited high stability at pH values from 7.0 to 10.0. The hydrolysis of β-cyclodextrin (β-CD) produced malto-oligosaccharides of various lengths. In addition to hydrolysis, MAG1 also demonstrated transglycosylation activity for the synthesis of longer malto-oligosaccharides. The thermodynamic equilibrium of the multiple reactions was shifted towards synthesis when the reaction conditions were optimised and the water activity was suppressed, which resulted in a yield of 38% transglycosylation products consisting of malto-oligosaccharides of various lengths. Thin layer chromatography and high-performance liquid chromatography analyses revealed the presence of malto-oligosaccharides with a higher degree of polymerisation than maltoheptaose, which has never been reported for other maltogenic amylases. The addition of organic solvents into the reaction further suppressed the water activity. The increase in the transglycosylation-to-hydrolysis ratio from 1.29 to 2.15 and the increased specificity toward maltopentaose production demonstrated the enhanced synthetic property of the enzyme. The high transglycosylation activity of maltogenic amylase offers a great advantage for synthesising malto-oligosaccharides and rare carbohydrates. PMID:25221964

  8. Chitosan oligosaccharides suppress production of nitric oxide in lipopolysaccharide-induced N9 murine microglial cells in vitro.

    PubMed

    Wei, Peng; Ma, Pan; Xu, Qing-Song; Bai, Qun-Hua; Gu, Jian-Guo; Xi, Hao; Du, Yu-Guang; Yu, Chao

    2012-08-01

    Chitosan oligosaccharides (COS) have been reported to exert many biological activities, such as antioxidant, antitumor and anti-inflammatory effects. In the present study, we examined the effect of COS on nitric oxide (NO) production in LPS induced N9 microglial cells. Pretreatment with COS (50~200 μg/ml) could markedly inhibit NO production by suppressing inducible nitric oxide synthase (iNOS) expression in activated microglial cells. Signal transduction studies showed that COS remarkably inhibited LPS-induced phosphorylation of p38 MAPK and ERK1/2. COS pretreatment could also inhibit the activation of both nuclear factor-κB (NF-κB) and activator protein-1 (AP-1). In conclusion, our results suggest that COS could suppress the production of NO in LPS-induced N9 microglial cells, mediated by p38 MAPK and ERK1/2 pathways.

  9. Xylo-oligosaccharides and lactitol promote the growth of Bifidobacterium lactis and Lactobacillus species in pure cultures.

    PubMed

    Mäkeläinen, H; Saarinen, M; Stowell, J; Rautonen, N; Ouwehand, A C

    2010-06-01

    The current screening study aimed at identifying promising prebiotic and synbiotic candidates. The fermentation of xylo-oligosaccharides, xylan, galacto-oligosaccharide, fructo-oligosaccharide, polydextrose, lactitol, gentiobiose and pullulan was investigated in vitro. The ability of these established and potential prebiotic candidates to function as a sole carbon source for probiotic (Bifidobacterium and Lactobacillus), intestinal and potential pathogenic microbes (Eubacterium, Bacteroides, Clostridium, Escherichia coli, Salmonella, and Staphylococcus) was assessed in pure cultures. Xylo-oligosaccharides were fermented with high specificity by the tested Bifidobacterium lactis strains and lactitol by lactobacilli, whereas galacto-oligosaccharides, fructo-oligosaccharides and gentiobiose were utilised by a larger group of microbes. Xylan, polydextrose and pullulan were utilised to a limited extent by only a few of the tested microbes. The results of this screening study indicate that xylo-oligosaccharides and lactitol support the growth of a limited number of beneficial microbes in pure cultures. Such a high degree of specificity has not been previously reported for established prebiotics. Based on these results, the most promising prebiotics and synbiotic combinations can be selected for further testing.

  10. High yield production of monomer-free chitosan oligosaccharides by pepsin catalyzed hydrolysis of a high deacetylation degree chitosan.

    PubMed

    Roncal, Tomás; Oviedo, Alberto; López de Armentia, Iratxe; Fernández, Laura; Villarán, M Carmen

    2007-12-28

    The high molecular weight of chitosan, which results in a poor solubility at neutral pH values and high viscosity aqueous solutions, limits its potential uses in the fields of food, health and agriculture. However, most of these limitations are overcome by chitosan oligosaccharides obtained by enzymatic hydrolysis of the polymer. Several commercial enzymes with different original specificities were assayed for their ability to hydrolyze a 93% deacetylation degree chitosan and compared with a chitosanase. According to the patterns of viscosity decrease and reducing end formation, three enzymes--cellulase, pepsin and lipase A--were found to be particularly suitable for hydrolyzing chitosan at a level comparable to that achieved by chitosanase. Unlike the appreciable levels of both 2-amino-2-deoxy-D-glucose and 2-acetamido-2-deoxy-D-glucose monomers released from chitosan by the other enzymes after a 20h-hydrolysis (4.6-9.1% of the total product weight), no monomer could be detected following pepsin cleavage. As a result, pepsin produced a higher yield of chitosan oligosaccharides than the other enzymes: 52% versus as much as 46%, respectively. Low molecular weight chitosans accounted for the remaining 48% of hydrolysis products. The calculated average polymerization degree of the products released by pepsin was around 16 units after 20h of hydrolysis. This product pattern and yield are proposed to be related to the bond cleavage specificity of pepsin and the high deacetylation degree of chitosan used as substrate. The optimal reaction conditions for hydrolysis of chitosan by pepsin were 40 degrees C and pH 4.5, and an enzyme/substrate ratio of 1:100 (w/w) for reactions longer than 1h.

  11. Production of arabinoxylan-oligosaccharide mixtures of varying composition from rye bran by a combination of process conditions and type of xylanase.

    PubMed

    Falck, Peter; Aronsson, Anna; Grey, Carl; Stålbrand, Henrik; Nordberg Karlsson, Eva; Adlercreutz, Patrick

    2014-12-01

    The aim was to study arabinoxylan-oligosaccharide production from rye bran using heat pretreatment and enzymatic hydrolysis. Due to the potential application in foods, the purity of arabinoxylan was also assessed. Rye bran was heat pretreated to improve xylanase-catalyzed hydrolysis of arabinoxylan into arabinoxylan-oligosaccharides. Enzymatic removal of starch and proteins before or after heat pretreatment increased the purity, although at lower yield. The most attractive process resulted in 62% (w/w) arabinoxylan content after ethanol precipitation. Using xylanases from two glycoside hydrolase families (RmXyn10A from GH10 and Pentopan Mono BG from GH11), different mixtures of unsubstituted and arabinose-substituted xylooligosaccharides were produced. GH10 gave a higher yield of short oligosaccharides (60%w/w) with xylobiose as the main product; xylobiose and xylotriose were the main products with GH11 (40%w/w). Thus, heat pretreatment combined with enzymatic hydrolysis can be used to produce arabinoxylan-oligosaccharides from rye bran that are potentially useful in functional foods. Copyright © 2014 Elsevier Ltd. All rights reserved.

  12. Purification and Characterization of Chitosanase from Bacillus sp. Strain KCTC 0377BP and Its Application for the Production of Chitosan Oligosaccharides

    PubMed Central

    Choi, Yeon Jin; Kim, Eun Jung; Piao, Zhe; Yun, Young Chul; Shin, Yong Chul

    2004-01-01

    For the enzymatic production of chitosan oligosaccharides from chitosan, a chitosanase-producing bacterium, Bacillus sp. strain KCTC 0377BP, was isolated from soil. The bacterium constitutively produced chitosanase in a culture medium without chitosan as an inducer. The production of chitosanase was increased from 1.2 U/ml in a minimal chitosan medium to 100 U/ml by optimizing the culture conditions. The chitosanase was purified from a culture supernatant by using CM-Toyopearl column chromatography and a Superose 12HR column for fast-performance liquid chromatography and was characterized according to its enzyme properties. The molecular mass of the enzyme was estimated to be 45 kDa by means of sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The enzyme demonstrated bifunctional chitosanase-glucanase activities, although it showed very low glucanase activity, with less than 3% of the chitosanase activity. Activity of the enzyme increased with an increase of the degrees of deacetylation (DDA) of the chitosan substrate. However, the enzyme still retained 72% of its relative activity toward the 39% DDA of chitosan, compared with the activity of the 94% DDA of chitosan. The enzyme produced chitosan oligosaccharides from chitosan, ranging mainly from chitotriose to chitooctaose. By controlling the reaction time and by monitoring the reaction products with gel filtration high-performance liquid chromatography, chitosan oligosaccharides with a desired oligosaccharide content and composition were obtained. In addition, the enzyme was efficiently used for the production of low-molecular-weight chitosan and highly acetylated chitosan oligosaccharides. A gene (csn45) encoding chitosanase was cloned, sequenced, and compared with other functionally related genes. The deduced amino acid sequence of csn45 was dissimilar to those of the classical chitosanase belonging to glycoside hydrolase family 46 but was similar to glucanases classified with glycoside hydrolase

  13. The potential for oligosaccharide production from the hemicellulose fraction of biomasses through pretreatment processes: xylooligosaccharides (XOS), arabinooligosaccharides (AOS), and mannooligosaccharides (MOS).

    PubMed

    Otieno, Daniel Obed; Ahring, Birgitte Kiaer

    2012-10-01

    Hemicellulosic oligosaccharides are sugar molecules that contain xylose, mannose, and arabinose in variable concentrations ranging from 3 to 10 molecules. These medium and long chain sugars can be classified as non-digestible carbohydrates, thus playing an important role in gastrointestinal health as prebiotics. Their physiological benefits, primarily stimulation of the proliferation of lactic acid bacteria and bifidobacteria in the colon informs their significance as high value nutraceuticals in the food and pharmaceutical industry. In addition they are well known as useful components of important pharmaceutical products. There are two main ways of producing these sugars from biomass, which include enzymatic and non-enzymatic pretreatments. Each of the two processes has advantages and disadvantages. Enzymatic processes are associated with high costs, higher concentration of monomeric sugars, and low oligosaccharide yields while thermo-chemical processes are usually associated with undesirable byproducts such as furfural and lower oligosaccharide yields. In this paper we discuss the benefits and constraints for optimization of different methods for the production of oligosaccharides from biomass. Copyright © 2012 Elsevier Ltd. All rights reserved.

  14. Assessment of suitability of vine shoots for hemicellulosic oligosaccharides production through aqueous processing.

    PubMed

    Dávila, Izaskun; Gordobil, Oihana; Labidi, Jalel; Gullón, Patricia

    2016-07-01

    Vine shoots were subjected to non-isothermal aqueous processing. A range of severities (S0) from 3.20 to 4.65 was assayed and their effects in terms of solubilization, composition, molar mass distribution, structural characterization and thermal stability of the liquors were studied using HPLC, HPSEC, TGA and FTIR. The spent solids were characterized by HPLC and FTIR. When autohydrolysis was carried out at S0=4.01, the substrate solubilization achieved a 38.7% of the raw material and 83.1% of the initial xylan was converted into xylooligosaccharides (XOS). The amount of TOS (total oligosaccharides) in the hydrolysates was 28.4g/L while the other non volatile compounds (ONVC) were 0.08g/g NVC. The spent solid from the treatment at S0=4.01 was composed about 90% of cellulose and lignin. Therefore, it can be concluded that autohydrolysis is a suitable pretreatment of vine shoots such as a first stage of a biomass refinery.

  15. Efficient Production of Prebiotic Gluco-oligosaccharides in Orange Juice Using Immobilized and Co-immobilized Dextransucrase.

    PubMed

    Tingirikari, Jagan Mohan Rao; Gomes, Wesley Faria; Rodrigues, Sueli

    2017-05-06

    Dextransucrase from Leuconostoc mesenteroides NRRL B-512F was subjected to immobilization and co-immobilization with dextranase from Chaetomium erraticum. Immobilization has enhanced the operational and storage stability of dextransucrase. Two hundred milligrammes (2.4 IU/mg) of alginate beads (immobilized and co-immobilized) were found to be optimum for the production of gluco-oligosaccharides (GOS) in orange juice with a high degree of polymerization. The pulp of the orange juice did not interfere in the reaction. In the batch process, co-immobilized dextransucrase (41 g/L) produced a significantly higher amount of GOS than immobilized dextransucrase (37 g/L). Alginate entrapment enhanced the thermal stability of dextransucrase for up to 3 days in orange juice at 30 °C. The production of GOS in semi-continuous process was 39 g/L in co-immobilized dextransucrase and 33 g/L in immobilized dextransucrase. Thus, immobilization technology offers a great scope in terms of reusability and efficient production of a value added functional health drink.

  16. Ultrasonication and steam-explosion as chitin pretreatments for chitin oligosaccharide production by chitinases of Lecanicillium lecanii.

    PubMed

    Villa-Lerma, Guadalupe; González-Márquez, Humberto; Gimeno, Miquel; López-Luna, Alberto; Bárzana, Eduardo; Shirai, Keiko

    2013-10-01

    In this study, chitin oligosaccharides have been successfully produced using chitinases from submerged fermentation of Lecanicillium lecanii. The highest Hex, Chit and Prot production was 0.14, 0.26 and 2.05 U/mg of protein, respectively, which were attained varying pH from 5 to 8 after 96 h. Culture conditions conducted at constant pH of 6 resulted in significantly lower enzyme production. The crude enzyme was partially purified by salting out with (NH4)2SO4 followed by size exclusion chromatography to isolate the chitinase mixture for further chitin hydrolysis assays. In this regard, chitin substrates were pretreated with sonication and steam explosion prior to enzymatic reaction. Structural changes were observed with steam explosion with 11.28% reduction of the crystallinity index attained with the lowest chitin/water ratio (0.1g/mL). Pretreated chitins reached the highest production of reducing sugars (0.37 mg/mL) and GlcNAc (0.59 mg/mL) in 23.6% yield.

  17. Purification and characterization of Vibrio parahaemolyticus extracellular chitinase and chitin oligosaccharide deacetylase involved in the production of heterodisaccharide from chitin.

    PubMed

    Kadokura, K; Rokutani, A; Yamamoto, M; Ikegami, T; Sugita, H; Itoi, S; Hakamata, W; Oku, T; Nishio, T

    2007-05-01

    A chitin-degrading bacterial strain, KN1699, isolated from Yatsu dry beach (Narashino, Chiba Prefecture, Japan), was identified as Vibrio parahaemolyticus. Treatment of powdered chitin with crude enzyme solution prepared from the supernatant of KN1699 cultures yielded a disaccharide, beta-D-N-acetylglucosaminyl-(1,4)-D-glucosamine (GlcNAc-GlcN), as the primary chitin degradation product. The extracellular enzymes involved in the production of this heterodisaccharide, chitinase (Pa-Chi; molecular mass, 92 kDa) and chitin oligosaccharide deacetylase (Pa-COD; molecular mass, 46 kDa), were isolated from the crude enzyme solution, and their hydrolysis specificities were elucidated. These studies confirmed that (1) Pa-Chi hydrolyzes chitin to produce (GlcNAc)(2) and (2) Pa-COD hydrolyzes the acetamide group of reducing end GlcNAc residue of (GlcNAc)(2). These findings indicate that GlcNAc-GlcN is produced from chitin by the cooperative hydrolytic reactions of both Pa-Chi and Pa-COD.

  18. Extraction of soluble arabinoxylan from enzymatically pretreated wheat bran and production of short xylo-oligosaccharides and arabinoxylo-oligosaccharides from arabinoxylan by glycoside hydrolase family 10 and 11 endoxylanases.

    PubMed

    Mathew, Sindhu; Karlsson, Eva Nordberg; Adlercreutz, Patrick

    2017-09-14

    The enzymatic, ecofriendly pretreatment of wheat bran with α-amylase from Bacillus amyloliquifaciens or B. licheniformis at 90°C for 1.5h followed by Neutrase at 50°C for 4h, aqueous liquefaction at 121°C for 15h and ethanol precipitation enabled the production of soluble arabinoxylan (AX) with purity of 70.9% and 68.4% (w/w) respectively. Process alternatives tried, to simplify the process and curtail the cost resulted in AX products with different purities, yields and arabinose to xylose ratio (A/X). Among the two glycoside hydrolase (GH) family endoxylanases evaluated, GH10 family hydrolysed soluble AX more efficiently with xylanase from Geobacillus stearothermophilus T-6 (GsXyn10A) producing maximum amount of quantifiable short xylo-oligosaccharides (XOS) and arabinoxylo-oligosaccharides (AXOS) (53% w/w) followed by the catalytic module of Rhodothermus marinus Xyn10A (RmXyn10A-CM) with 37% (w/w) conversion. The GH11 family endoxylanases, from Thermomyces lanuginosus (Pentopan Mono BG™) and Neocallimastix patriciarum (NpXyn11A) gave conversions of 21% and 22% (w/w) of the soluble AX, respectively (major AXOS products were not quantified). In addition to the XOS formed such as X2, X3 and X4, the AXOS products identified were A(3)X and A(2)XX in the case of GsXyn10A and RmXyn10A-CM while Pentopan Mono BG and NpXyn11A produced XA(3)XX as the major AXOS product. Copyright © 2017 Elsevier B.V. All rights reserved.

  19. The mixture of bifidobacterium associated with fructo-oligosaccharides reduces the damage of the ocular surface.

    PubMed

    Chisari, G; Chisari, E M; Francaviglia, A; Chisari, C G

    2017-01-01

    Despite its high prevalence Dry Eye Syndrome (DES) in frequently under-recognized owing to its negative influence on patients visual function. This clinical trial was a pilot study to evaluate the effects of supplementation with mixture (Bifidobacterium lactis and Bifidobacterium bifido) on the tear film. Following the run-in period subjects were randomized in two groups: group A (N°20 subjects) and group B (N°20 subjects). Group A (control) treated only with substitute tear and group B treated with substitute tear + mixture (symbiotic). The data obtained in the two study groups A and B were, respectively the following: Schirmer 9.1±0.2 vs 12.7±0.4 (p< 0.001); Schirmer II 3.5±0.1 VS 4.7±0.2 (p<0.001); BUT 3.9±0.3 vs 6.3±0.2 (p<0.001). Culture test showed initial bacterial growth in group "A" (placebo) 18 out of 40 samples tested, corresponding to 45.0% and "B" after treatment ((symbiotic) was found positive culture whit growth of bacteria in 12 tests equal to 30.0%. The total numbers of isolations of aerobic and anaerobic bacteria found group A and B after treatment. A reduction of 15 to 11 strains of aerobic and anaerobic isolates from 9 to 5 has been found. The present study shows that the administration of bifidobacterium may represent a success full treatment in ameliorating dry eye syndrome (DES). The effect of imbalanced microbiota are not restricted by gastrointestinal abnormalities but could have systemic impact on immunity. Commensal bacteria or probiotics interact with the endogenous enteric microbiota and gut cells therein confereing health benefit to the host.

  20. Hemicellulose isolation, characterization, and the production of xylo-oligosaccharides from the wastewater of a viscose fiber mill.

    PubMed

    Zhang, Yuedong; Yu, Guang; Li, Bin; Mu, Xindong; Peng, Hui; Wang, Haisong

    2016-05-05

    Viscose fiber mills generate a lot of wastewater enriched with hemicelluloses. The structure of the hemicellulose in the wastewater was characterized and the hemicellulose was isolated to produce xylo-oligosaccharides (XOS). It was confirmed that the hemicellulose was mainly 4-O-methylglucuronoxylan with a small amount of glucomannan and xyloglucan. The 4-O-methylglucuronoxylan was completely de-acetylated and linear with a few 4-O-methyl glucuronic acid attached. After purified by the acid precipitation and washing, the hemicellulose was pretreated by dilute acid, and then subjected to xylanase hydrolysis. After the dilute H2SO4 pretreatment at pH 2.6 and 150°C for 30min and the followed xylanase hydrolysis (65IU/g xylan), the total XOS yield was improved from 0.215 to 0.578g/g xylan. The percentage of XOS in the final sugar product was 68.9%. These results demonstrated the potential economical and environmental benefits of the process to utilize the byproducts from viscose fiber mills.

  1. Cellulosimicrobium cellulans strain E4-5 enzymatic hydrolysis of curdlan for production of (1 → 3)-linked β-D-glucan oligosaccharides.

    PubMed

    Fu, Yunbin; Cheng, Likun; Meng, Yanyu; Li, Shuguang; Zhao, Xiaoming; Du, Yuguang; Yin, Heng

    2015-12-10

    In order to find an efficient enzymatic tool for curdlan degradation to produce (1 → 3)-linked β-D-glucan oligosaccharides, strain E4-5 (registration number JN089883, Genbank) was isolated from seaside soil. The 16S rRNA gene sequencing classified it as Cellulosimicrobium cellulans. It was the first reported microorganism that succeeded in degrading high-set heated curdlan blocks. The ferments of strain E4-5 also showed good degradation effects on laminaran and alkali-neutralized curdlan. Due to the products with less amount of glucose, it was assumed that endo-1,3-β-glucanases of strain E4-5 had a greater hydrolyzing effect than exo-1,3-β-glucanases. This indicated that strain E4-5 was a promising microorganism to hydrolyze (1 → 3)-linked β-D-glucan. Moreover, alkali-neutralization pretreatment was effective for promoting a more diversified degree of polymerization (DP) of (1 → 3)-linked β-D-glucan oligosaccharides under enzymatic hydrolysis and will pave the way for making full use of curdlan for production of glucan oligosaccharides.

  2. Purification and functional characterization of endo-beta-mannanase MAN5 and its application in oligosaccharide production from konjac flour.

    PubMed

    Zhang, Min; Chen, Xiu-Lan; Zhang, Zhi-Hua; Sun, Cai-Yun; Chen, Lei-Lei; He, Hai-Lun; Zhou, Bai-Cheng; Zhang, Yu-Zhong

    2009-07-01

    MAN5, the main extracellular saccharide hydrolase from Bacillus sp. MSJ-5, is an endo-beta-mannanase with a demand of at least five sugar moieties for effective cleavage. It has a pH optimum of 5.5 and a temperature optimum of 50 degrees C and is stable at pH 5-9 or below 65 degrees C. MAN5 has a very high ability to hydrolyze konjac flour, 10 U/mg of which could completely liquefy konjac flour gum in 10 min at 50 degrees C. HPLC analysis showed that most glucomannan in the konjac flour was hydrolyzed into a large amount of oligosaccharides with DP of 2-6 and a very small amount of monosaccharide. With the culture supernatant as enzyme source, the optimum condition to prepare oligosaccharides from konjac flour was obtained as 10 mg/ml konjac flour incubated with 10 U/mg enzyme at 50 degrees C for 24 h. With this condition, more than 90% polysaccharides in the konjac flour solution were hydrolyzed into oligosaccharides and a little monosaccharide (2.98% of the oligosaccharides). Konjac flour is an underutilized agricultural material with low commercial value in China. With MAN5, konjac flour can be utilized to generate high value-added oligosaccharides. The high effectiveness and cheapness of this technique indicates its potential in industry.

  3. Production of high-purity isomalto-oligosaccharides syrup by the enzymatic conversion of transglucosidase and fermentation of yeast cells.

    PubMed

    Pan, Yen-Chung; Lee, Wen-Chien

    2005-03-30

    A method for the production of high-purity isomalto-oligosaccharides (IMO) involving the transglucosylation by transglucosidase and yeast fermentation was proposed. The starch of rice crumbs was enzymatically liquefied and saccharified, and then converted to low-purity IMO syrup by transglucosylation. The low-purity IMO produced either from rice crumbs or tapioca flour as the starch source could be effectively converted to high-purity IMO by yeast fermentation to remove the digestible sugars including glucose, maltose, and maltotriose. Both Saccharomyces carlsbergensis and Saccharomyces cerevisiae were able to ferment glucose in the IMO syrup. Cells of S. carlsbergensis harvested from the medium of malt juice were also able to ferment maltose and maltotriose. A combination of these two yeasts or S. carlsbergensis alone could be used to totally remove the digestible sugars in the IMO, coupled with the production of ethanol. The resultant high-purity IMO, including mainly isomaltose, panose, and isomaltotriose made up more than 98% w/w of the total sugars after a 3-day fermentation. When the low-purity IMO was produced from the starch of tapioca flour, 3-day fermentation under the same conditions resulted in IMO with purity lower than that from rice crumbs. For low-purity IMO from rice crumbs, fermentation with washed S. carlsbergensis cells harvested at log phase was the most effective. However, for the low-purity IMO from tapioca flour, incubation with S. cerevisiae for the first 24 h and then supplementing with an equal amount of S. carlsbergensis cells for further fermentation was the most effective approach for producing high-purity IMO.

  4. Nature and biosynthesis of galacto-oligosaccharides related to oligosaccharides in human breast milk

    PubMed Central

    Intanon, Montira; Arreola, Sheryl Lozel; Pham, Ngoc Hung; Kneifel, Wolfgang; Haltrich, Dietmar; Nguyen, Thu-Ha

    2014-01-01

    Human milk oligosaccharides (HMO) are prominent among the functional components of human breast milk. While HMO have potential applications in both infants and adults, this potential is limited by the difficulties in manufacturing these complex structures. Consequently, functional alternatives such as galacto-oligosaccharides are under investigation, and nowadays, infant formulae are supplemented with galacto-oligosaccharides to mimic the biological effects of HMO. Recently, approaches toward the production of defined human milk oligosaccharide structures using microbial, fermentative methods employing single, appropriately engineered microorganisms were introduced. Furthermore, galactose-containing hetero-oligosaccharides have attracted an increasing amount of attention because they are structurally more closely related to HMO. The synthesis of these novel oligosaccharides, which resemble the core of HMO, is of great interest for applications in the food industry. PMID:24571717

  5. Alteration of membrane oligosaccharides by castanospermine, an alpha glucosidase inhibitor, enhances immunoglobulin production in Staphylococcus aureus Cowan I-stimulated lymphocyte culture.

    PubMed

    Karasuno, T; Kanayama, Y; Nishiura, T; Nakao, H; Kurata, Y; Yonezawa, T; Tarui, S

    1990-11-01

    Castanospermine (CSP) inhibits alpha-glucosidase, which is involved in the initial step of N-linked oligosaccharide processing of secretory and membrane glycoproteins. In Staphylococcus aureus Cowan I (SAC)-stimulated human lymphocyte culture, CSP at a dose of 20 micrograms/ml caused a twofold increase in immunoglobulin G (IgG) release after 7 days. An initial 48-h exposure to CSP sufficed for this enhancing effect. Plaque-forming cell assays on the seventh day disclosed that CSP caused an increase in the number of IgG-, IgA- and IgM-secreting cells. In cross-culture experiments, only a mixture of B cells pretreated with CSP and untreated T cells showed an increase in IgG production. Tritiated thymidine incorporation studies revealed that CSP enhanced B-cell responses to T cell-derived soluble factor (TSF). When incubated with CSP for 18 h, B cells showed an increased surface binding on [3H]concanavalin A (Con A). These results indicate that the alteration in B-cell membrane oligosaccharides enhances the response to TSF at an early stage of SAC culture, leading to an increase in Ig-secreting cell number at later stages. The present study provides evidence that cell-surface oligosaccharides of B cells play an important role in the responses of B cells to lymphokines.

  6. Strategic glycan elution map for the production of human-type N-linked oligosaccharides: the case of hen egg yolk and white.

    PubMed

    Sumiyoshi, Wataru; Nakakita, Shin-ichi; Miyanishi, Nobumitsu; Hirabayashi, Jun

    2009-03-23

    Glycans play important roles in various biological phenomena, but the lack of a systematic procedure for producing complex structures of glycans severely restricts their application in the medical and industrial fields. In this paper, we propose a basic strategy for the preparation of substantial amounts (>100 mg) of N-linked oligosaccharides, where the structure of each glycan is mapped with its elution position in liquid chromatography as well as the empirical yield. In model experiments using hen egg white and yolk as starting materials, the former provided a series of agalactosylated complex-type and hybrid-type N-linked oligosaccharides containing bisecting N-acetylglucosamine (GlcNAc) in addition to two high-mannose type glycans. In contrast, egg yolk gave predominantly alpha2-6sialylated biantennary glycans together with a high-mannose type one, reflecting the difference in the origins of the tissues. Due to the total identity of the glycans obtained to human ones, the present strategy should provide a practical scheme for the production of human-type N-linked oligosaccharides.

  7. A new effective process for production of curdlan oligosaccharides based on alkali-neutralization treatment and acid hydrolysis of curdlan particles in water suspension.

    PubMed

    Li, Jing; Zhu, Li; Zheng, Zhi-Yong; Zhan, Xiao-Bei; Lin, Chi-Chung; Zong, Yu; Li, Wei-Jiang

    2013-10-01

    Biologically active β-1,3-oligosaccharides with rapidly growing biomedical applications are produced from hydrolysis of curdlan polysaccharide. The water-insoluble curdlan impedes its hydrolysis efficiency which is enhanced by our newly developed alkali-neutralization treatment process to increase the stability of curdlan suspension to more than 20 days, while the untreated control settled within 5 min. A putative double-layer structure model comprising of a compact core and a hydrated outer layer was proposed to describe the treated curdlan particles based on sedimentation and scanning electron microscopy observation. This model was verified by single- and two-step acid hydrolysis, indicative of the reduced susceptibility to hydrolysis when close to the compact core. Electrospray ionization-mass spectrometry, thin-layer chromatography analyses, and effective HPLC procedure led to the development of improved process to produce purified individual β-1,3-oligosaccharides with degrees of polymerization from 2 to 10 and potential for biomedical applications from curdlan hydrolyzate. Our new curdlan oligosaccharide production process offers an even better alternative to the previously published processes.

  8. Dietary-fiber-degrading enzymes from a human intestinal Clostridium and their application to oligosaccharide production from nonstarchy polysaccharides using immobilized cells.

    PubMed

    Nakajima, N; Ishihara, K; Matsuura, Y

    2002-07-01

    The secretion of nonstarchy polysaccharide-degrading enzymes from an anaerobic human intestinal bacterium, Clostridium butyricum- beijerinckii (isolated from human feces), was investigated. Growth of the bacterium was found when laminarin, konjac glucomannan, and pectic acid were added separately to the culture media as sole carbon source. The corresponding degrading enzymes for these dietary fibers, laminarinase (endo-1,3- beta-glucanase), endo-1,4-beta-mannanase, endo- and exo-pectate lyases, and pectin methylesterase, were then purified and characterized. These extracelluar enzymes, which were secreted by the bacterium in the human large intestine, were considered to contribute to digestion of the ingested dietary fibers to their oligosaccharides, following by short-chain fatty acid fermentation by the bacterium. We have developed cell immobilization techniques of the bacterium on cellulose-foam carriers that are effective for continuous production of the oligosaccharides from the dietary fibers in a fed-batch reactor system. From 9 g of pectic acid, a total of 3.96 g of 4,5-unsaturated digalacturonic acid was produced over 40 h in four 500-ml batchcultures. In the same manner, the corresponding oligosaccharides were obtained from konjac glucomannan and laminarin with average conversion rates of around 30-40%.

  9. Human milk and related oligosaccharides as prebiotics.

    PubMed

    Barile, Daniela; Rastall, Robert A

    2013-04-01

    Human milk oligosaccharides (HMO) are believed to have a range of biological activities beyond providing nutrition to the infant. Principal among these is that they may act as prebiotics. Prebiotics are dietary ingredients, usually oligosaccharides that provide a health benefit to the host mediated by the modulation of the human gut microbiota. While it is clear that such oligosaccharides may have potential applications in infants and adults alike, this potential is limited by the difficulties in manufacturing HMO. Consequently functional alternatives such as galacto-oligosaccharides (GOS) are under investigation. GOS are produced enzymatically from lactose for commercial use in food applications--including addition to infant formulae--as similar to breast milk oligosaccharides, they encourage a gut bacteria population that promotes health and reduces the incidence of intestinal infections. New methods for separation and concentration of complex, breast milk-like oligosaccharides from bovine milk industrial streams that contain only low amounts of these valuable oligosaccharides are providing the opportunity to investigate other viable sources of specific oligosaccharides for use as prebiotics in supplements or food products.

  10. Bifidobacterium fermented milk and galacto-oligosaccharides lead to improved skin health by decreasing phenols production by gut microbiota.

    PubMed

    Miyazaki, K; Masuoka, N; Kano, M; Iizuka, R

    2014-06-01

    A questionnaire survey found that women suffering from abnormal bowel movements have many skin problems such as a high frequency of dry skin. Although there are similarities between the structure and barrier function mechanism of the gut and skin, experimental data are insufficient to show an association between the intestinal environment and skin conditions. Phenols, for example phenol and p-cresol, as metabolites of aromatic amino acids produced by gut bacteria, are regarded as bioactive toxins and serum biomarkers of a disturbed gut environment. Recent studies have demonstrated that phenols disturb the differentiation of monolayer-cultured keratinocytes in vitro, and that phenols produced by gut bacteria accumulate in the skin via the circulation and disrupt keratinocyte differentiation in hairless mice. Human studies have demonstrated that restriction of probiotics elevated serum free p-cresol levels and harmed skin conditions (reduced skin hydration, disrupted keratinisation). In contrast, daily intake of the prebiotic galacto-oligosaccharides (GOS) restored serum free p-cresol levels and skin conditions in adult women. Moreover, a double-blind placebo-controlled trial demonstrated that the daily intake of fermented milk containing the probiotic Bifidobacterium breve strain Yakult and prebiotic GOS reduced serum total phenol levels and prevented skin dryness and disruption of keratinisation in healthy adult women. It is concluded that phenols produced by gut bacteria are one of the causes of skin problems. Probiotics and/or prebiotics, such as B. breve strain Yakult and/or GOS, are expected to help maintain a healthy skin by decreasing phenols production by gut microbiota. These findings support the hypothesis that probiotics and prebiotics provide health benefits to the skin as well as the gut.

  11. Comparison of Xylo-oligosaccharides production by autohydrolysis of fibers separated from ground corn flour and DDGS

    USDA-ARS?s Scientific Manuscript database

    Xylo-oligosaccharides (XOS) are known to have beneficial health properties, and are considered to be functional food ingredients. The objective of this study is to compare corn fibers separated from ground corn flour and distillers dried grains with solubles (DDGS) for XOS yield and optimum authoyd...

  12. Characterization of a Novel Fructosyltransferase from Lactobacillus reuteri That Synthesizes High-Molecular-Weight Inulin and Inulin Oligosaccharides

    PubMed Central

    van Hijum, S. A. F. T.; van Geel-Schutten, G. H.; Rahaoui, H.; van der Maarel, M. J. E. C.; Dijkhuizen, L.

    2002-01-01

    Fructosyltransferase (FTF) enzymes produce fructose polymers (fructans) from sucrose. Here, we report the isolation and characterization of an FTF-encoding gene from Lactobacillus reuteri strain 121. A C-terminally truncated version of the ftf gene was successfully expressed in Escherichia coli. When incubated with sucrose, the purified recombinant FTF enzyme produced large amounts of fructo-oligosaccharides (FOS) with β-(2→1)-linked fructosyl units, plus a high-molecular-weight fructan polymer (>107) with β-(2→1) linkages (an inulin). FOS, but not inulin, was found in supernatants of L. reuteri strain 121 cultures grown on medium containing sucrose. Bacterial inulin production has been reported for only Streptococcus mutans strains. FOS production has been reported for a few bacterial strains. This paper reports the first-time isolation and molecular characterization of (i) a Lactobacillus ftf gene, (ii) an inulosucrase associated with a generally regarded as safe bacterium, (iii) an FTF enzyme synthesizing both a high molecular weight inulin and FOS, and (iv) an FTF protein containing a cell wall-anchoring LPXTG motif. The biological relevance and potential health benefits of an inulosucrase associated with an L. reuteri strain remain to be established. PMID:12200292

  13. Study on preparation and separation of Konjac oligosaccharides.

    PubMed

    Jian, Wenjie; Sun, Yuanming; Huang, Huan; Yang, Youhui; Peng, Shuhui; Xiong, Bo; Pan, Tingtiao; Xu, Zhenlin; He, Mingxiang; Pang, Jie

    2013-02-15

    To study the preparation and separation of Konjac oligosaccharides, Konjac Glucomannan was degraded by the combination of γ-irradiation and β-mannanase, and then the degradation product was separated by ultrafiltration. To our interest, for most of Konjac oligosaccharides obtained by this method, the molecular mass was lower than 2200 Da. In addition, the 1000 Da molecular weight cut off membrane could effectively separate the Konjac oligosaccharides. In conclusion, the combination of γ-irradiation and β-mannanase was an efficient method to obtain Konjac oligosaccharides, and the oligosaccharides of molecular mass lower than 1000 Da could be effectively separated by ultrafiltration. Copyright © 2012 Elsevier Ltd. All rights reserved.

  14. In Vitro Fermentation of Sugar Beet Arabino-Oligosaccharides by Fecal Microbiota Obtained from Patients with Ulcerative Colitis To Selectively Stimulate the Growth of Bifidobacterium spp. and Lactobacillus spp. ▿ †

    PubMed Central

    Vigsnæs, Louise Kristine; Holck, Jesper; Meyer, Anne S.; Licht, Tine Rask

    2011-01-01

    The potential prebiotic properties of arabino-oligosaccharides (AOS) derived from sugar beet pulp was studied using mixed cultures of human fecal bacteria from patients with ulcerative colitis (UC), in remission or with active disease, and in healthy controls. These results were compared to those for fructo-oligosaccharides (FOS), which are known to have a prebiotic effect. Fermentation studies were carried out using a small-scale static batch system, and changes in the fecal microbial communities and metabolites were monitored after 24 h by quantitative real-time PCR and short-chain fatty acid analysis. With a few minor exceptions, AOS affected the communities similarly to what was seen for FOS. Quantitative real-time PCR revealed that Bifidobacterium spp. and Lactobacillus spp. were selectively increased after fermentation of AOS or FOS by fecal microbiota derived from UC patients. The stimulation of growth of Lactobacillus spp. and Bifidobacterium spp. was accompanied by a high production of acetate and hence a decrease of pH. The fermentation of AOS may help improve the inflammatory conditions in UC patients through stimulation of bacteria eliciting anti-inflammatory responses and through production of acetate. AOS may therefore represent a new prebiotic candidate for reduction of the risk of flare-ups in UC patients. However, human trials are needed to confirm a health-promoting effect. PMID:21984234

  15. In vitro fermentation of sugar beet arabino-oligosaccharides by fecal microbiota obtained from patients with ulcerative colitis to selectively stimulate the growth of Bifidobacterium spp. and Lactobacillus spp.

    PubMed

    Vigsnæs, Louise Kristine; Holck, Jesper; Meyer, Anne S; Licht, Tine Rask

    2011-12-01

    The potential prebiotic properties of arabino-oligosaccharides (AOS) derived from sugar beet pulp was studied using mixed cultures of human fecal bacteria from patients with ulcerative colitis (UC), in remission or with active disease, and in healthy controls. These results were compared to those for fructo-oligosaccharides (FOS), which are known to have a prebiotic effect. Fermentation studies were carried out using a small-scale static batch system, and changes in the fecal microbial communities and metabolites were monitored after 24 h by quantitative real-time PCR and short-chain fatty acid analysis. With a few minor exceptions, AOS affected the communities similarly to what was seen for FOS. Quantitative real-time PCR revealed that Bifidobacterium spp. and Lactobacillus spp. were selectively increased after fermentation of AOS or FOS by fecal microbiota derived from UC patients. The stimulation of growth of Lactobacillus spp. and Bifidobacterium spp. was accompanied by a high production of acetate and hence a decrease of pH. The fermentation of AOS may help improve the inflammatory conditions in UC patients through stimulation of bacteria eliciting anti-inflammatory responses and through production of acetate. AOS may therefore represent a new prebiotic candidate for reduction of the risk of flare-ups in UC patients. However, human trials are needed to confirm a health-promoting effect.

  16. Extensive dry ball milling of wheat and rye bran leads to in situ production of arabinoxylan oligosaccharides through nanoscale fragmentation.

    PubMed

    Van Craeyveld, Valerie; Holopainen, Ulla; Selinheimo, Emilia; Poutanen, Kaisa; Delcour, Jan A; Courtin, Christophe M

    2009-09-23

    This study investigated the potential of ball milling as a dry process for in situ production of arabinoxylan oligosaccharides (AXOS) in arabinoxylan (AX)-rich wheat and rye bran. An extensive lab-scale ball mill treatment (120 h, 50% jar volume capacity) increased the wheat bran water-extractable arabinoxylan (WE-AX) level from 4% (untreated bran) to 61% of the wheat bran AX. Extractable AX fragments had an arabinose/xylose ratio (A/X ratio) of 0.72 and a molecular mass (MM) of 15 kDa. Ball milling of rye bran gave rise to similar results, with the A/X ratio of the extractable AX being considerably lower (0.51). Optimization of the ball mill treatment by varying the degree of filling of the milling jar permitted us to obtain equally high WE-AX levels (>70%) in wheat and rye bran within a 24 h ball-milling period. Ball milling at optimal conditions (24 h, 16% jar volume capacity) yielded wheat bran AXOS, with an A/X ratio of 0.65 and a MM of 6 kDa. Ball milling (24 h, 50% jar volume capacity) of pericarp-enriched wheat bran increased the WE-AX level from 1% (untreated pericarp) to 63%. The extractable material had a high A/X ratio (0.97) and a low MM (5 kDa). Fluorescence microscopy revealed that the extensive ball mill treatment led to the almost complete disappearance of discernible tissue structures in the ball-milled material, indicating bran particle size reductions down to the nanoscale level. It further visualized the aggregation of the treated material. These results show that AXOS can be produced in situ from wheat or rye bran in a single-stage dry milling process, rendering a wet extraction step redundant. The higher A/X ratio of the obtained AXOS than of enzymically produced wheat bran-derived AXOS offers perspectives for the production of a wide range of AXOS structures. Moreover, ball milling makes upgrading of the low-value pericarp layer feasible.

  17. Sucrose and Related Oligosaccharides

    NASA Astrophysics Data System (ADS)

    Eggleston, Gillian

    Sucrose (α-D-glucopyranosyl-(1↔2)-β-D-fructofuranoside) is the most common low-molecular-weight sugar found in the plant kingdom. It is ubiquitously known as common table sugar and primarily produced industrially from sugarcane (Saccharum officinarum) and sugar beet (Beta vulgaris); the basics of the industrial manufacture of sucrose are outlined in this chapter. Commercial sucrose has a very high purity (> 99.9%) making it one of the purest organic substances produced on an industrial scale. Value-addition to sucrose via chemical and biotechnological reactions is becoming more important for the diversification of the sugar industry to maintain the industries' competitiveness in a world increasingly turning to a bio-based economy. The basis for the chemical reactivity of sucrose is the eight hydroxyl groups present on the molecule, although, sucrose chemical reactivity is regarded as difficult. Increasing use of enzymatic biotechnological techniques to derivatize sucrose is expected, to add special functionalities to sucrose products like biodegradability, biocompatibility, and non-toxicity. Analysis of sucrose by colorimetric, enzymatic, oxidation-reduction and chromatography methods are discussed. Oligosaccharides related to sucrose are outlined in detail and include sucrose-based plant, honey and in vitro oligosaccharides.

  18. Adsorption and kinetic behavior of recombinant multifunctional xylanase in hydrolysis of pineapple stem and bagasse and their hemicellulose for Xylo-oligosaccharide production.

    PubMed

    Zhao, Li-Chao; Wang, Yan; Lin, Jun-Fang; Guo, Li-Qiong

    2012-04-01

    A recombinant multifunctional xylanase (RMFXase) was utilized to hydrolyze xylan in pineapple stems and bagasse. The feasibility of using RMFXase and the two agricultural by-products for producing Xylo-oligosaccharides was demonstrated. Adsorption dynamics showed that lignin and cellulose delayed the adsorption equilibrium. The adsorption data fit the Langmuir-type adsorption isotherm. The extent of hydrolysis was quantified by measuring the reducing and total sugars in substrate-RMFXase complexes at 50°C. The substrates had similar hydrolysis characteristics. Differences in the maximum hydrolysis rate (HR) and the time of maximum were obtained. The HR(max) of water-insoluble oat spelt xylan reached 55.26% after 2h. Analysis of the end-products using high-performance liquid chromatography indicated xylobiose and xylotriose were the major hydrolysis products, and the final degree of polymerization was greater than two. The purity of the substrates and RMFXase are the key factors that determine the processing parameters of Xylo-oligosaccharide production. Copyright © 2012 Elsevier Ltd. All rights reserved.

  19. Variations in prebiotic oligosaccharide fermentation by intestinal lactic acid bacteria.

    PubMed

    Endo, Akihito; Nakamura, Saki; Konishi, Kenta; Nakagawa, Junichi; Tochio, Takumi

    2016-01-01

    Prebiotic oligosaccharides confer health benefits on the host by modulating the gut microbiota. Intestinal lactic acid bacteria (LAB) are potential targets of prebiotics; however, the metabolism of oligosaccharides by LAB has not been fully characterized. Here, we studied the metabolism of eight oligosaccharides by 19 strains of intestinal LAB. Among the eight oligosaccharides used, 1-kestose, lactosucrose and galactooligosaccharides (GOSs) led to the greatest increases in the numbers of the strains tested. However, mono- and disaccharides accounted for more than half of the GOSs used, and several strains only metabolized the mono- and di-saccharides in GOSs. End product profiles indicated that the amounts of lactate produced were generally consistent with the bacterial growth recorded. Oligosaccharide profiling revealed the interesting metabolic manner in Lactobacillus paracasei strains, which metabolized all oligosaccharides, but left sucrose when cultured with fructooligosaccharides. The present study clearly indicated that the prebiotic potential of each oligosaccharide differs.

  20. Production of a recombinant chitin oligosaccharide deacetylase from Vibrio parahaemolyticus in the culture medium of Escherichia coli cells.

    PubMed

    Kadokura, Kazunari; Sakamoto, Yusuke; Saito, Kaori; Ikegami, Takanori; Hirano, Takako; Hakamata, Wataru; Oku, Tadatake; Nishio, Toshiyuki

    2007-08-01

    An open reading frame (ORF) encoding chitin oligosaccharide deacetylase (Pa-COD) gene and its signal sequence was cloned from the Vibrio parahaemolyticus KN1699 genome and its sequence was analyzed. The ORF encoded a 427 amino acid protein, including the 22 amino acid signal sequence. The deduced amino acid sequence was highly similar to several bacterial chitin oligosaccharide deacetylases in carbohydrate esterase family 4. An expression plasmid containing the gene was constructed and inserted into Escherichia coli cells and the recombinant enzyme was secreted into the culture medium with the aid of the signal peptide. The concentration of the recombinant enzyme in the E. coli culture medium was 150 times larger than that of wild-type enzyme produced in the culture medium by V. parahaemolyticus KN1699. The recombinant enzyme was purified to homogeneity from culture supernatant in an overall yield of 16%. Substrate specificities of the wild-type and the recombinant enzymes were comparable.

  1. Enhanced Production of κ-Carrageenase and κ-Carrageenan Oligosaccharides through Immobilization of Thalassospira sp. Fjfst-332 with Magnetic Fe3O4-Chitosan Microspheres.

    PubMed

    Guo, Juanjuan; Zheng, Zhichang; Chen, Chi; Lu, Xu; Zhang, Yi; Zheng, Baodong

    2017-09-13

    In this study, immobilized bacteria (IMB) microsphere was prepared by embedding κ-carrageenase-producing Thalassospira sp. Fjfst-332 (TF332) onto a magnetic Fe3O4-chitosan carrier. The performance of Fe3O4-chitosan carrier was optimized by comparing its bacteria immobilization capacity at different Fe3O4:chitosan ratios and temperatures, while the functions of IMB microspheres were characterized by examining their κ-carrageenase production at different temperatures, pH's, and reuse cycles. At the 1:1 (w:w) Fe3O4:chitosan ratio, the Fe3O4-chitosan carriers possessed sufficient anchoring capacity for bacterial immobilization without significant compromise of their magnetism for magnetic separation of IMB from culture media. The spectroscopic analysis of IMB microspheres indicated that the immobilization of TF332 might affect the amide groups in chitosan. Compared to free bacteria, IMB can produce κ-carrageenase at higher temperature, wider pH range, and faster rate. More importantly, the κ-carrageenase-producing activity was sustained for at least seven reuse cycles. The major κ-carrageenan degradation products of IMB-derived κ-carrageenase were the oligosaccharides containing two to six monosaccharide units. Overall, this Fe3O4-chitosan-TF-332 microsphere has the potential to become a stable and reusable platform for large-scale production of κ-carrageenan oligosaccharides.

  2. Non-digestible oligosaccharides used as prebiotic agents: mode of production and beneficial effects on animal and human health.

    PubMed

    Grizard, D; Barthomeuf, C

    1999-01-01

    Prebiotic agents are food ingredients that are potentially beneficial to the health of consumers. The main commercial prebiotic agents consist of oligosaccharides and dietary fibres (mainly inulin). They are essentially obtained by one of three processes: 1) the direct extraction of natural polysaccharides from plants; 2) the controlled hydrolysis of such natural polysaccharides; 3) enzymatic synthesis, using hydrolases and/or glycosyl transferases. Both of these enzyme types catalyse transglycosylation reactions, allowing synthesis of small molecular weight synthetic oligosaccharides from mono- and disaccharides. Presently, in Europe, inulin-type fructans, characterised by the presence of fructosyl units bound to the beta-2,1 position of sucrose, are considered as one of the carbohydrate prebiotic references. Prebiotics escape enzymatic digestion in the upper gastrointestinal tract and enter the caecum without change to their structure. None are excreted in the stools, indicating that they are fermented by colonic flora so as to give a mixture of short-chain fatty acids (acetate, propionate and butyrate), L-lactate, carbon dioxide and hydrogen. By stimulating bifidobacteria, they may have the following implications for health: 1) potential protective effects against colorectal cancer and infectious bowel diseases by inhibiting putrefactive bacteria (Clostridium perfringens ) and pathogen bacteria (Escherichia coli, Salmonella, Listeria and Shigella ), respectively; 2) improvement of glucid and lipid metabolisms; 3) fibre-like properties by decreasing the renal nitrogen excretion; 4) improvement in the bioavailability of essential minerals; and 5) low cariogenic factor. These potential beneficial effects have been largely studied in animals but have not really been proven in humans. The development of a second generation of oligosaccharides and the putative implication of a complex bacterial trophic chain in the intestinal prebiotic fermentation process are also

  3. Change in oligosaccharides during processing of soybean sheet.

    PubMed

    Wang, Qiushuang; Ke, Leqin; Yang, Dongmei; Bao, Bili; Jiang, Jianmei; Ying, Tiejin

    2007-01-01

    Oligosaccharides have been credited with many health-promoting functions, which had been identified in many clinical studies, such as promoting the growth of Bifidobacterium in human intestine and balance of intestinal bacteria, modulating the immune response, inhibition of cancer and tumor, stimulation of mineral absorption. In this study the effect of processing unit operations on the levels of soybean oligosaccharides during production of soybean sheet were investigated. The concentrations of oligosaccharide in initial raw soybean were: sucrose 43.05 g/kg, raffinose 7.52 g/kg and stachyose 41.32 g/kg (in dry matter). Oligosaccharide losses in the soaking water, in the first filtrating stage, in the second filtrating stage and finally in the sheet formation stage were 0.68, 10.3, 8.15 and 47.22 g/kg (initial dry soybean) respectively, representing 0.74, 11.21, 8.87 and 51.39% of the total oligosaccharides present in the initial soybeans. The recovery of oligosaccharides in the final soybean sheet from the initial soybean was 27.92%. The loss of soybean oligosaccharides in different processing stages, especially in the by-product, the sweet slurry, was considerable. The loss of oligosaccharides was mainly associated with water/matter removal in production process. The analysis of loss profile implied possible ways to improve the technology for production of oligosaccharides-enriched soy-sheets.

  4. Enzymatic synthesis of prebiotic oligosaccharides.

    PubMed

    Rabelo, Maria C; Honorato, Talita L; Gonçalves, Luciana R B; Pinto, Gustavo A S; Rodrigues, Sueli

    2006-04-01

    Prebiotic oligosaccharides are nondigestible carbohydrates that can be obtained by enzymatic synthesis. Glucosyltransferases can be used to produce these carbohydrates through an acceptor reaction synthesis. When maltose is the acceptor a trisaccharide composed of one maltose unit and one glucose unit linked by an alpha-1,6-glycosidic bond (panose) is obtained as the primer product of the dextransucrase acceptor reaction. In this work, panose enzymatic synthesis was evaluated by a central composite experimental design in which maltose and sucrose concentration were varied in a wide range of maltose/sucrose ratios in a batch reactor system. A partially purified enzyme was used in order to reduce the process costs, because enzyme purification is one of the most expensive steps in enzymatic synthesis. Even using high maltose/sucrose ratios, dextran and higher-oligosaccharide formation were not avoided. The results showed that intermediate concentrations of sucrose and high maltose concentration resulted in high panose productivity with low dextran and higher-oligosaccharide productivity.

  5. Arrival time distributions of product ions reveal isomeric ratio of deprotonated molecules in ion mobility-mass spectrometry of hyaluronan-derived oligosaccharides.

    PubMed

    Hermannová, Martina; Iordache, Andreea-Maria; Slováková, Kristína; Havlíček, Vladimír; Pelantová, Helena; Lemr, Karel

    2015-06-01

    Hyaluronic acid is a naturally occurring linear polysaccharide with substantial medical potential. In this work, discrimination of tyramine-based hyaluronan derivatives was accessed by ion mobility-mass spectrometry of deprotonated molecules and nuclear magnetic resonance spectroscopy. As the product ion mass spectra did not allow for direct isomer discrimination in mixture, the reductive labeling of oligosaccharides as well as stable isotope labeling was performed. The ion mobility separation of parent ions together with the characteristic fragmentation for reduced isomers providing unique product ions allowed us to identify isomers present in a mixture and determine their mutual isomeric ratio. The determination used simple recalculation of arrival time distribution areas of unique ions to areas of deprotonated molecules. Mass spectrometry data were confirmed by nuclear magnetic resonance spectroscopy.

  6. Preweaning modulation of intestinal microbiota by oligosaccharides or amoxicillin can contribute to programming of adult microbiota in rats.

    PubMed

    Morel, Fanny B; Oozeer, Raish; Piloquet, Hugues; Moyon, Thomas; Pagniez, Anthony; Knol, Jan; Darmaun, Dominique; Michel, Catherine

    2015-03-01

    Increasing evidence suggests that early nutrition has programming effects on adult health. Identifying mechanisms underlying nutritional programming would aid in the design of new disease prevention strategies. The intestinal microbiota could be a key player in this programming because it affects host metabolic homeostasis, postnatal gut colonization is sensitive to early nutrition, and initial microbial set-up is thought to shape microbiota composition for life. The aim of this study was to determine whether early manipulation of intestinal microbiota actually programs adult microbiota in rats. Suckling rats pups were supplemented with fructo-oligosaccharides, galacto-oligosaccharides/long-chain fructan mix (GOS/lcF, 9/1), acidic oligosaccharides, amoxicillin, or vehicle from the fifth to the fourteenth day of life, and weaned to standard chow at day 21. Ceco-colonic microbiota was characterized at 14 and 131 d by real-time polymerase chain reaction analysis. At day 14, all treatments affected microbiota. Amoxicillin had the most significant effect. All oligosaccharides decreased Firmicutes levels, whereas only fructo-oligosaccharides and GOS/lcF increased bifidobacteria. At day 131, most of these effects had faded away but a significant, albeit minor, adult microbiota programming was observed for rats that received GOS/lcF mix before weaning, regarding Roseburia intestinalis cluster, one subdivision of the Erysipelotrichaceae family as well as butyrate kinase gene. As revealed by a targeted quantitative polymerase chain reaction approach, programming of adult intestinal microbiota seems to vary according to the nature of the preweaning microbiotal modulator. This suggests that intestinal microbiota may, only under specific circumstances, serve as a relay of neonatal nutrition and thus potentially contribute to nutritional programming of host physiology. Copyright © 2015 Elsevier Inc. All rights reserved.

  7. Effects of dietary oligosaccharide supplementation on growth performance, concentrations of the major odor-causing compounds in excreta, and the cecal microflora of broilers.

    PubMed

    Yang, G Q; Yin, Y; Liu, H Y; Liu, G H

    2016-10-01

    This study investigated the effects of dietary supplementation with 4 types of oligosaccharides on the growth performance, concentrations of the major odor-causing compounds in excreta and cecal microflora of broilers. Three hundred 21-day-old Archer Abor broilers with an average initial live weight of 702.3 g were randomly divided into 5 dietary treatments: basal diet, basal diet + 5 g/kg of mannan-oligosaccharide (MOS), basal diet + 1.2 g/kg of inulin, basal diet + 1.5 g/kg of fructo-oligosaccharide (FOS), and basal diet +1.25 g/kg of soybean oligosaccharide (SBOS), respectively. Each diet was fed to 6 replicates of 10 birds from d 21 to 42, and body weight and feed intake were recorded. Fresh excreta were sampled from each replicate on d 40, 41, and 42 and stored at -20 °C until further analysis. On d 42, the ceca of killed birds were aseptically removed, and the cecal contents were collected into sterile containers and stored at -80 °C until further analysis. Results showed that feeding inulin, FOS, and SBOS diets resulted in an improvement in daily gain (P < 0.05). Broilers fed the SBOS diet showed lower feed:gain ratio (1.84g:g) than the other groups (P > 0.05). Broilers fed the FOS diet showed the lowest volatile basic nitrogen, pH value, and indole and skatole contents in excreta, and broilers fed the SBOS diet had higher total volatile fatty acids concentrations than control (P < 0.05). The cecal microbial community was measured using the PCR-DGGE, which indicated that the cecal microflora Shannon-wiener index and richness of SBOS-fed broilers were significantly higher than that of the control (P < 0.05). The lowest evenness was recorded in the FOS group, which was significantly lower than the other groups (P < 0.05) except the SBOS group. Based on the sequences of the corresponding 16S rDNA of the DGGE bands, in combination with the contents of the major odor-causing compounds in excreta, it is suggested that uncultured Lachnospiraceae bacterium

  8. In vivo measurement of flatulence and nutrient digestibility in dogs fed poultry by-product meal, conventional soybean meal, and low-oligosaccharide low-phytate soybean meal.

    PubMed

    Yamka, Ryan M; Harmon, David L; Schoenherr, William D; Khoo, Christina; Gross, Kathy L; Davidson, Stephen J; Joshi, Dinesh K

    2006-01-01

    To determine an optimal window for determining peak flatulence and evaluate the effects of oligosaccharides and supplemental beta-mannanase in soybean meal-based diets on nutrient availability and flatulence. 6 dogs. Dogs were used in a 2 x 3 factorial arrangement of treatments in a 6 x 6 Latin square experiment to evaluate the digestibility, flatulence, and fecal odor metabolites of low-oligosaccharide low-phytate soybean meal (LLM), conventional soybean meal (SBM), and poultry by-product (PBP) meal diets with or without supplemental beta-mannanase (5 g/kg). Enzyme supplementation had no effect on total tract dry matter (DM), nitrogen digestibility, or digestible energy; however, differences between protein sources did exist for total tract DM digestibility and digestible energy. The PBP meal had higher DM digestibility and digestible energy (mean, 0.913 and 4,255 cal/g), compared with soy-based diets (mean, 0.870 and 4,049 cal/g). No differences were detected for any treatment regardless of protein source or addition of supplemental enzyme for any flatulence components analyzed. No differences were detected for all fecal odor metabolites regardless of addition of supplemental enzyme; however, differences between protein sources were detected. The PBP meal had lower concentrations of carboxylic acids and esters and higher concentrations of heterocycles, phenols, thio and sulfides, ketones, alcohols, and indoles than LLM and SBM. Diets containing < 22.4 g of stachyose/kg and < 2 g of raffinose/kg did not alter digestibility or increase flatulence in dogs.

  9. Oligosaccharides in feces of breast- and formula-fed babies.

    PubMed

    Albrecht, Simone; Schols, Henk A; van Zoeren, Diny; van Lingen, Richard A; Groot Jebbink, Liesbeth J M; van den Heuvel, Ellen G H M; Voragen, Alphons G J; Gruppen, Harry

    2011-10-18

    So far, little is known on the fate of oligosaccharides in the colon of breast- and formula-fed babies. Using capillary electrophoresis with laser induced fluorescence detector coupled to a mass spectrometer (CE-LIF-MS(n)), we studied the fecal oligosaccharide profiles of 27 two-month-old breast-, formula- and mixed-fed preterm babies. The interpretation of the complex oligosaccharide profiles was facilitated by beforehand clustering the CE-LIF data points by agglomerative hierarchical clustering (AHC). In the feces of breast-fed babies, characteristic human milk oligosaccharide (HMO) profiles, showing genetic fingerprints known for human milk of secretors and non-secretors, were recognized. Alternatively, advanced degradation and bioconversion of HMOs, resulting in an accumulation of acidic HMOs or HMO bioconversion products was observed. Independent of the prebiotic supplementation of the formula with galactooligosaccharides (GOS) at the level used, similar oligosaccharide profiles of low peak abundance were obtained for formula-fed babies. Feeding influences the presence of diet-related oligosaccharides in baby feces and gastrointestinal adaptation plays an important role herein. Four fecal oligosaccharides, characterized as HexNAc-Hex-Hex, Hex-[Fuc]-HexNAc-Hex, HexNAc-[Fuc]-Hex-Hex and HexNAc-[Fuc]-Hex-HexNAc-Hex-Hex, highlighted an active gastrointestinal metabolization of the feeding-related oligosaccharides. Their presence was linked to the gastrointestinal mucus layer and the blood-group determinant oligosaccharides therein, which are characteristic for the host's genotype.

  10. Bacterial metabolism and health-related effects of galacto-oligosaccharides and other prebiotics.

    PubMed

    Macfarlane, G T; Steed, H; Macfarlane, S

    2008-02-01

    Most studies involving prebiotic oligosaccharides have been carried out using inulin and its fructo-oligosaccharide (FOS) derivatives, together with various forms of galacto-oligosaccharides (GOS). Although many intestinal bacteria are able to grow on these carbohydrates, most investigations have demonstrated that the growth of bifidobacteria, and to a lesser degree lactobacilli, is particularly favoured. Because of their safety, stability, organoleptic properties, resistance to digestion in the upper bowel and fermentability in the colon, as well as their abilities to promote the growth of beneficial bacteria in the gut, these prebiotics are being increasingly incorporated into the Western diet. Inulin-derived oligosaccharides and GOS are mildly laxative, but can result in flatulence and osmotic diarrhoea if taken in large amounts. However, their effects on large bowel habit are relatively minor. Although the literature dealing with the health significance of prebiotics is not as extensive as that concerning probiotics, considerable evidence has accrued showing that consumption of GOS and FOS can have significant health benefits, particularly in relation to their putative anti-cancer properties, influence on mineral absorption, lipid metabolism, and anti-inflammatory and other immune effects such as atopic disease. In many instances, prebiotics seem to be more effective when used as part of a synbiotic combination.

  11. Intestinal microbiology in early life: specific prebiotics can have similar functionalities as human-milk oligosaccharides.

    PubMed

    Oozeer, Raish; van Limpt, Kees; Ludwig, Thomas; Ben Amor, Kaouther; Martin, Rocio; Wind, Richèle D; Boehm, Günther; Knol, Jan

    2013-08-01

    Human milk is generally accepted as the best nutrition for newborns and has been shown to support the optimal growth and development of infants. On the basis of scientific insights from human-milk research, a specific mixture of nondigestible oligosaccharides has been developed, with the aim to improve the intestinal microbiota in early life. The mixture has been extensively studied and has been shown to be safe and to have potential health benefits that are similar to those of human milk. The specific mixture of short-chain galacto-oligosaccharides and long-chain fructo-oligosaccharides has been found to affect the development of early microbiota and to increase the Bifidobacterium amounts as observed in human-milk-fed infants. The resulting gut ecophysiology is characterized by high concentrations of lactate, a slightly acidic pH, and specific short-chain fatty acid profiles, which are high in acetate and low in butyrate and propionate. Here, we have summarized the main findings of dietary interventions with these specific oligosaccharides on the gut microbiota in early life. The gut ecophysiology in early life may have consequences for the metabolic, immunologic, and even neurologic development of the child because reports increasingly substantiate the important function of gut microbes in human health. This review highlights major findings in the field of early gut colonization and the potential impact of early nutrition in healthy growth and development.

  12. Heterodisaccharide 4-O-(N-acetyl-beta-D-glucosaminyl)-D-glucosamine is a specific inducer of chitinolytic enzyme production in Vibrios harboring chitin oligosaccharide deacetylase genes.

    PubMed

    Hirano, Takako; Kadokura, Kazunari; Ikegami, Takanori; Shigeta, Yuko; Kumaki, Yasuko; Hakamata, Wataru; Oku, Tadatake; Nishio, Toshiyuki

    2009-09-01

    Vibrio parahaemolyticus KN1699 produces 4-O-(N-acetyl-beta-d-glucosaminyl)-d-glucosamine (GlcNAc-GlcN) as a major end product from chitin using two extracellular hydrolases: glycoside hydrolase family 18 chitinase, which produces (GlcNAc)(2) from chitin, and carbohydrate esterase (CE) family 4 chitin oligosaccharide deacetylase (COD), which hydrolyzes the N-acetyl group at the reducing-end GlcNAc residue of (GlcNAc)(2). In this study, we clarified that this heterodisaccharide functions as an inducer of the production of the two above-mentioned chitinolytic enzymes, particularly chitinase. Similar results for chitinase production were obtained with other chitin-decomposing Vibrio strains harboring the CE family 4 COD gene; however, such an increase in chitinase production was not observed in chitinolytic Vibrio strains that did not harbor the COD gene. These results suggest that GlcNAc-GlcN is a unique inducer of chitinase production in Vibrio bacteria that have the COD-producing ability and that the COD involved in the synthesis of this signal compound is one of the key enzymes in the chitin catabolic cascade of these bacteria.

  13. Production of High Galacto-oligosaccharides by Pectinex Ultra SP-L: Optimization of Reaction Conditions and Immobilization on Glyoxyl-Functionalized Silica.

    PubMed

    González-Delgado, Isabel; Segura, Yolanda; Morales, Gabriel; López-Muñoz, María-José

    2017-03-01

    A rational optimization for the synthesis of galacto-oligosaccharides (GOS) from lactose catalyzed by β-galactosidase from Aspergillus aculeatus, included in the commercial product Pectinex Ultra SP-L, has been performed by using experimental design and surface response methodology. This accurate tool optimized empirical production of the most desired high-GOS (tri-GOS and tetra-GOS) up to 16.4% under the following reaction conditions: 59 °C, 4 U/mL free enzyme concentration, pH 6.5, 250 g/L initial lactose concentration, and 20 h of reaction. The statistical analysis revealed temperature and initial lactose concentration as critical parameters. The successful immobilization of the enzyme on a glyoxyl-functionalized porous silica support slightly increased the yield toward high-GOS (17.6%), especially tri-GOS yield (15.3%), under the optimized reaction conditions as compared to the free enzyme. Furthermore, the promotion of the transgalactosylation reaction toward tri-GOS production increased 1.5-fold the productivity of high-GOS as compared to the free enzyme.

  14. Anaphylaxis to cow's milk formula containing short-chain galacto-oligosaccharide.

    PubMed

    Chiang, Wen Chin; Huang, Chiung-Hui; Llanora, Genevieve V; Gerez, Irvin; Goh, Si Hui; Shek, Lynette P C; Nauta, Alma J; Van Doorn, Wilhelmina A; Bindels, Jacques; Ulfman, Laurien H; Knipping, Karen; Delsing, Dianne J; Knol, Edward F; Lee, Bee Wah

    2012-12-01

    On the basis of the proven prebiotic effects of oligosaccharides in cow's milk formula (CMF) in infants, CMFs are supplemented with oligosaccharides. We present a series of 5 cases of cow's milk-tolerant but atopic patients with a history of respiratory allergies. All had anaphylaxis after the ingestion of CMF supplemented with short-chain galacto-oligosaccharide (scGOS). The allergen trigger was investigated. Clinical histories were collated. Skin prick tests (SPTs) and basophil activation tests (BATs) were carried out with the eliciting CMF that triggered anaphylaxis, with or without supplemented prebiotics (scGOS) and with scGOS fractions containing oligosaccharides of different chain lengths. The median age of presentation was 6 years (range, 5-38 years). Anaphylaxis occurred within 30 minutes of the first known exposure to CMF supplemented with prebiotics in all patients. Only 1 patient was subjected to oral challenge, which resulted in an anaphylactic reaction. All patients demonstrated IgE sensitization through SPTs and BATs to scGOS and fractions of scGOS containing 3 sugar units or greater but not to cow's milk or long-chain fructo-oligosaccharide. Eight child control subjects tolerant to regular ingestion of scGOS-supplemented CMF and 1 adult volunteer were found to have negative results to scGOS through SPTs and BATs. In addition, in vitro BATs with donor basophils sensitized with sera from 2 of the 3 reported cases showed reactions to scGOS. The scGOS-induced basophil activation was inhibited in the presence of wortmannin, a phosphatidylinositol 3-kinase inhibitor. This study describes an unusual form of IgE-mediated anaphylaxis triggered by low-molecular-weight oligosaccharides in scGOS. The primary sensitizer for this phenomenon requires further investigation. Copyright © 2012 American Academy of Allergy, Asthma & Immunology. Published by Mosby, Inc. All rights reserved.

  15. Immobilization of xylanase from Bacillus pumilus strain MK001 and its application in production of xylo-oligosaccharides.

    PubMed

    Kapoor, Mukesh; Kuhad, Ramesh Chander

    2007-08-01

    Xylanase from Bacillus pumilus strain MK001 was immobilized on different matrices following varied immobilization methods. Entrapment using gelatin (GE) (40.0%), physical adsorption on chitin (CH) (35.0%), ionic binding with Q-sepharose (Q-S) (45.0%), and covalent binding with HP-20 beads (42.0%) showed the maximum xylanase immobilization efficiency. The optimum pH of immobilized xylanase shifted up to 1.0 unit (pH 7.0) as compared to free enzyme (pH 6.0). The immobilized xylanase exhibited higher pH stability (up to 28.0%) in the alkaline pH range (7.0-10.0) as compared to free enzyme. Optimum temperature of immobilized xylanase was observed to be 8 degrees C higher (68.0 degrees C) than free enzyme (60.0 degrees C). The free xylanase retained 50.0% activity, whereas xylanase immobilized on HP-20, Q-S, CH, and GE retained 68.0, 64.0, 58.0, and 57.0% residual activity, respectively, after 3 h of incubation at 80.0 degrees C. The immobilized xylanase registered marginal increase and decrease in Km and Vmax values, respectively, as compared to free enzyme. The immobilized xylanase retained up to 70.0% of its initial hydrolysis activity after seven enzyme reaction cycles. The immobilized xylanase was found to produce higher levels of high-quality xylo-oligosaccharides from birchwood xylan, indicating its potential in the nutraceutical industry.

  16. Prebiotic oligosaccharides in premature infants.

    PubMed

    Underwood, Mark A; Kalanetra, Karen M; Bokulich, Nicholas A; Mirmiran, Majid; Barile, Daniela; Tancredi, Daniel J; German, J Bruce; Lebrilla, Carlito B; Mills, David A

    2014-03-01

    The aim of the study was to determine the impact of increasing doses of 2 prebiotic oligosaccharides and of an "all-human diet" on the intestinal microbiota of premature infants. Twelve premature infants receiving formula feedings were randomly assigned to receive either galacto-oligosaccharide (F+GOS) or a pooled concentrated donor human milk product containing human milk oligosaccharides (F+HMO) in increasing doses during a 5-week period. A second group of 15 premature infants received their mother's own milk fortified with either a concentrated donor human milk product (H+H) or a bovine powdered fortifier (H+B). Serial stool specimens from each infant were analyzed by terminal restriction fragment length polymorphism and quantitative polymerase chain reaction for bacterial composition. All of the infants studied had relatively low levels of bifidobacteria and no measurable Lactobacilli. Infants from the F+GOS and F+HMO groups demonstrated an increase in relative numbers of Clostridia with increasing doses. Compared with the H+B group, the infants in the F+HMO and the H+H groups showed an unexpected trend toward an increase in γ-Proteobacteria over time/dose. Principal coordinate analyses and Shannon diversity scores were not significantly different among the 4 groups. Infants in the H+H group received more antibiotics during the study period than those in the other groups. Two of the infants receiving GOS developed feeding intolerance. None of the prebiotic interventions resulted in significant increases in bifidobacteria compared with baseline specimens or the H+B group; however, many of the infants did not receive the highest doses of GOS and HMO, and antibiotic use in the H+H group was high.

  17. Oligosaccharides in infant formula: more evidence to validate the role of prebiotics.

    PubMed

    Vandenplas, Yvan; Zakharova, Irina; Dmitrieva, Yulia

    2015-05-14

    The gastrointestinal (GI) microbiota differs between breast-fed and classic infant formula-fed infants. Breast milk is rich in prebiotic oligosaccharides (OS) and may also contain some probiotics, but scientific societies do not recommend the addition of prebiotic OS or probiotics to standard infant formula. Nevertheless, many infant formula companies often add one or the other or both. Different types of prebiotic OS are used in infant formula, including galacto-oligosaccharide, fructo-oligosaccharide, polydextrose and mixtures of these OS, but none adds human milk OS. There is evidence that the addition of prebiotics to infant formula brings the GI microbiota of formula-fed infants closer to that of breast-fed infants. Prebiotics change gut metabolic activity (by decreasing stool pH and increasing SCFA), have a bifidogenic effect and bring stool consistency and defecation frequency closer to those of breast-fed infants. Although there is only limited evidence that these changes in GI microbiota induce a significant clinical benefit for the immune system, interesting positive trends have been observed in some markers. Additionally, adverse effects are extremely seldom. Prebiotics are added to infant formula because breast milk contains human milk OS. Because most studies suggest a trend of beneficial effects and because these ingredients are very safe, prebiotics bring infant formula one step closer to the golden standard of breast milk.

  18. Experimental studies and two-dimensional modelling of a packed bed bioreactor used for production of galacto-oligosaccharides (GOS) from milk whey.

    PubMed

    Sen, Pramita; Bhattacharjee, Chiranjib; Bhattacharya, Pinaki

    2016-03-01

    In the present study, extensive experimental investigations and detailed theoretical analysis of a two-dimensional packed bed bioreactor, employed for the production of galacto-oligosaccharides (GOS) from milk whey were performed. Model equations, in one- and two-dimensions, capable of predicting the substrate concentration distribution in the bioreactor were developed by coupling mass balance equation with appropriate velocity distribution equation and solved numerically. Validation of the proposed model equations was done by a set of experimental data obtained from the bioreactor. The effects of reactor to catalyst particle diameter ratio (d t/d p), feed flowrate (10(-6)-10(-9) m(3) s(-1)), and initial lactose concentration (50-200 kg m(-3)) on substrate concentration distribution were investigated in detail. While, the distribution of substrate concentration in axial direction was independent of d t/d p, it was observed that for d t/d p <40, significant radial concentration distribution existed. It was further observed that the substrate conversion and product yield obtained experimentally showed an excellent agreement (97 ± 2 %) with the results predicted by the two-dimensional model equation, whereas, the results predicted by the one-dimensional model equation did not lie within the desired confidence level (<90 %). The results were confirmed by both curve fitting and statistical analysis. The prediction of substrate concentration distribution in axial and radial directions using the developed two-dimensional model equation is necessary for computing the bioreactor volume to achieve the desired GOS yield.

  19. INHIBITORY EFFECT OF CHITOSAN OLIGOSACCHARIDE ON HUMAN HEPATOMA CELLS IN VITRO.

    PubMed

    Liu, Likun; Xin, Yi; Liu, Jia; Zhang, Ershao; Li, Weiling

    2017-01-01

    Chitosan oligosaccharide, the degradation products of chitin, was reported to have a wide range of physiological functions and biological activities. In this study, we explored the inhibitory effect of Chitosan oligosaccharide on human hepatoma cells. MTT assay was applied to detect cell viability of the human hepatoma cells treated with Chitosan oligosaccharide. Flow cytometric analysis was used to investigate the apoptosis of the human hepatoma cells treated with Chitosan oligosaccharide. We employed western blot to investigate the underlying mechanisms involved in the apoptosis. Our data indicated that chitosan oligosaccharide dose-dependently inhibited the growth of hepatoma cells and induced apoptosis. On the molecular level, chitosan oligosaccharide decreased Bcl-2 and increased Caspase-3 expression which may be related to the apoptosis of hepatoma cells. Our results provide an experimental basis for the clinical development of Chitosan oligosaccharide as a novel anti-hepatoma drug.

  20. In vitro evaluation of the prebiotic activity of a pectic oligosaccharide-rich extract enzymatically derived from bergamot peel.

    PubMed

    Mandalari, G; Nueno Palop, C; Tuohy, K; Gibson, G R; Bennett, R N; Waldron, K W; Bisignano, G; Narbad, A; Faulds, C B

    2007-01-01

    The prebiotic effect of a pectic oligosaccharide-rich extract enzymatically derived from bergamot peel was studied using pure and mixed cultures of human faecal bacteria. This was compared to the prebiotic effect of fructo-oligosaccharides (FOS). Individual species of bifidobacteria and lactobacilli responded positively to the addition of the bergamot extract, which contained oligosaccharides in the range of three to seven. Fermentation studies were also carried out in controlled pH batch mixed human faecal cultures and changes in gut bacterial groups were monitored over 24 h by fluorescent in situ hybridisation, a culture-independent microbial assessment. Addition of the bergamot oligosaccharides (BOS) resulted in a high increase in the number of bifidobacteria and lactobacilli, whereas the clostridial population decreased. A prebiotic index (PI) was calculated for both FOS and BOS after 10 and 24 h incubation. Generally, higher PI scores were obtained after 10 h incubation, with BOS showing a greater value (6.90) than FOS (6.12).

  1. Rice bran polysaccharides and oligosaccharides modified by Grifola frondosa fermentation: Antioxidant activities and effects on the production of NO.

    PubMed

    Liu, Qian; Cao, Xiujuan; Zhuang, Xuhui; Han, Wei; Guo, Weiqun; Xiong, Jian; Zhang, Xiaolin

    2017-05-15

    Rice bran polysaccharides (RBPSs) are valuable compounds with many biological activities. In this work, a fungus called Grifola frondosa, was selected to ferment defatted rice bran water extracts and modify the RBPSs, which were then isolated by ethanol precipitation and deproteinization. GC analysis of fermented products suggested they are composed of glucose, arabinose, galactose, mannose, and xylose at a molar ratio of 9:5:8:2:5, which was 32:4:6:2:5 before fermentation. HPLC analysis revealed that the molecular weight of unfermented RBPS was distributed mainly from 10(3) to 10(4)Da, and it changed to 10(2) to 10(3)Da after fermentation. Antioxidant activities and effects on the production of NO were analyzed and it indicated that the scavenging ratios of hydroxyl and DPPH radicals by the fermented products were significantly enhanced compared to the unfermented ones, and also the products fermented for 9days exhibited two-way adjusting effects on the production of NO in macrophages. Copyright © 2016. Published by Elsevier Ltd.

  2. The relevance of tick bites to the production of IgE antibodies to the mammalian oligosaccharide galactose-α-1,3-galactose

    PubMed Central

    Commins, Scott P.; James, Hayley R.; Kelly, Elizabeth A.; Pochan, Shawna L.; Workman, Lisa J.; Perzanowski, Matthew S.; Kocan, Katherine M.; Fahy, John V.; Nganga, Lucy W.; Ronmark, Eva; Cooper, Philip J.; Platts-Mills, Thomas A. E.

    2011-01-01

    Background In 2009, we reported a novel form of delayed anaphylaxis to red meat, which is related to serum IgE antibodies to the oligosaccharide galactose-alpha-1,3-galactose (alpha-gal). Most of these patients had tolerated meat for many years previously. The implication is that some exposure in adult life had stimulated the production of these IgE antibodies. Objectives To investigate possible causes of this IgE antibody response, focusing on evidence related to tick bites, which are common in the region where these reactions occur. Methods Serum assays were carried out using biotinylated proteins and extracts bound to a streptavidin ImmunoCAP. Results Prospective studies on IgE antibodies in three subjects following tick bites showed an increase in IgE to alpha-gal of twenty-fold or greater. Other evidence included i) a strong correlation between histories of tick bites and IgE to alpha-gal (χ2=26.8, p<0.001), ii) evidence that these IgE antibodies are common in areas where the tick Amblyomma americanum is common, and iii) a significant correlation between IgE antibodies to alpha-gal and IgE antibodies to proteins derived from A. americanum (rs=0.75, p<0.001). Conclusion The results presented here provide evidence that tick bites are a cause, or possibly the only cause, of IgE specific for alpha-gal in this area of the United States. Both the number of subjects becoming sensitized and the titer of IgE antibodies to alpha-gal are striking. Here we report the first example of a response to an ectoparasite giving rise to an important form of food allergy. PMID:21453959

  3. Purification, physicochemical and thermodynamic studies of antifungal chitinase with production of bioactive chitosan-oligosaccharide from newly isolated Aspergillus griseoaurantiacus KX010988.

    PubMed

    Shehata, Abeer N; Abd El Aty, Abeer A; Darwish, Doaa A; Abdel Wahab, Walaa A; Mostafa, Faten A

    2017-09-20

    In our search for chitinase and chitosanase producer from unconventional sources, the marine-derived fungus Aspergillus griseoaurantiacus KX010988 was obviously the best producer of the highest chitinase and chitosanase activities by solid state fermentation of potato shells. Chitinase was purified in three steps involving ammonium sulphate precipitation, DEAE-cellulose ion-exchange chromatography and Sephacryl S-300 gel chromatography. 12.55 fold increase in purity with a recovery of 17.6 was obtained. The molecular mass of the purified chitinase was found to be 130kDa. It was optimally active at pH 4.5 and 40°C. Km and Vmax values were 0.22mgmL(-1) and 19.6μmolemin(-1)mg(-1) respectively. Mn(2+) and Zn(2+) ions lead to increased chitinase activity. While Fe(2+)and Cu(2+)ions strongly inhibited the chitinase activity. The thermodynamics of pure chitinase including activation energy for thermal denaturation (Ea,d), change of free energy (ΔGd), enthalpy(ΔHd), entropy(ΔSd) and half life values (T1/2) at 40, 50 and 60°C were determined. Chitinase showed antifungal activity against pathogenic fungus Fusarium solani. Chitosanase was partially purified by acetone precipitation (50-75%) v/v concentration. The hydrolytic products of moderate molecular weight of chitosan by chitosanase were analyzed by thin layer chromatography (TLC) after 12 and 24h respectively. Chitosan-oligosaccharides showed good antibacterial and antioxidant activities. Copyright © 2017. Published by Elsevier B.V.

  4. In Vitro Fermentation of Porcine Milk Oligosaccharides and Galacto-oligosaccharides Using Piglet Fecal Inoculum.

    PubMed

    Difilippo, Elisabetta; Pan, Feipeng; Logtenberg, Madelon; Willems, Rianne H A M; Braber, Saskia; Fink-Gremmels, Johanna; Schols, Henk A; Gruppen, Harry

    2016-03-16

    In this study, the in vitro fermentation by piglet fecal inoculum of galacto-oligosaccharides (GOS) and porcine milk oligosaccharides (PMOs) was investigated to identify possible preferences for individual oligosaccharide structures by piglet microbiota. First, acidic PMOs and GOS with degrees of polymerization 4-7 were depleted within 12 h of fermentation, whereas fucosylated and phosphorylated PMOs were partially resistant to fermentation. GOS structures containing β1-3 and β1-2 linkages were preferably fermented over GOS containing β1-4 and β1-6 linkages. Upon in vitro fermentation, acetate and butyrate were produced as the main organic acids. GOS fermentation by piglet inoculum showed a unique fermentation pattern with respect to preference of GOS size and organic acids production.

  5. Enzymatic saccharification of sugar beet pulp for the production of galacturonic acid and arabinose; a study on the impact of the formation of recalcitrant oligosaccharides.

    PubMed

    Leijdekkers, A G M; Bink, J P M; Geutjes, S; Schols, H A; Gruppen, H

    2013-01-01

    Enzymatic saccharification of sugar beet pulp was optimized on kg-scale to release the maximum amounts of monomeric galacturonic acid and arabinose with limited concomitant degradation of cellulose, using conditions that are feasible for industrial upscaling. A selected mixture of pectinases released 79% of the galacturonic acid and 82% of the arabinose as monomers from sugar beet pulp while simultaneously degrading only 17% of the cellulose. The recalcitrant structures that were obtained after hydrolysis were characterized using mass spectrometry. The most abundant structures had an average degree of polymerization of 4-5. They were identified as partially acetylated rhamnogalacturonan-oligosaccharides, mostly containing a terminal galacturonosyl residue on both reducing and non-reducing end, partially methyl esterified/acetylated homogalacturonan-oligosaccharides, mostly containing methyl and acetyl esters at contiguous galacturonosyl residues and arabinan-oligosaccharides, hypothesized to be mainly branched. It could be concluded that especially rhamnogalacturonan-galacturonohydrolase, arabinofuranosidase and pectin acetylesterase are lacking for further degradation of recalcitrant oligosaccharides.

  6. Trends in dairy and non-dairy probiotic products - a review.

    PubMed

    Vijaya Kumar, Bathal; Vijayendra, Sistla Venkata Naga; Reddy, Obulam Vijaya Sarathi

    2015-10-01

    Health awareness has grown to a greater extent among consumers and they are looking for healthy probiotic counterparts. Keeping in this view, the present review focuses recent developments in dairy and non-dairy probiotic products. All over the world, dairy probiotics are being commercialized in many different forms. However, the allergy and lactose intolerance are the major set-backs to dairy probiotics. Whereas, flavor and refreshing nature are the major advantages of non-dairy drinks, especially fruit juices. Phenotypic and genotypic similarities between dairy and non-dairy probiotics along with the matrix dependency of cell viability and cell functionality are reviewed. The heterogeneous food matrices of non-dairy food carriers are the major constraints for the survival of the probiotics, while the probiotic strains from non-dairy sources are satisfactory. Technological and functional properties, besides the viability of the probiotics used in fermented products of non-dairy origin are extremely important to get a competitive advantage in the world market. The functional attributes of dairy and non-dairy probiotic products are further enhanced by adding prebiotics such as galacto-oligosaccharide, fructo-oligosaccharide and inulin.

  7. Enzymatic production of defined chitosan oligomers with a specific pattern of acetylation using a combination of chitin oligosaccharide deacetylases

    PubMed Central

    Hamer, Stefanie Nicole; Cord-Landwehr, Stefan; Biarnés, Xevi; Planas, Antoni; Waegeman, Hendrik; Moerschbacher, Bruno Maria; Kolkenbrock, Stephan

    2015-01-01

    Chitin and chitosan oligomers have diverse biological activities with potentially valuable applications in fields like medicine, cosmetics, or agriculture. These properties may depend not only on the degrees of polymerization and acetylation, but also on a specific pattern of acetylation (PA) that cannot be controlled when the oligomers are produced by chemical hydrolysis. To determine the influence of the PA on the biological activities, defined chitosan oligomers in sufficient amounts are needed. Chitosan oligomers with specific PA can be produced by enzymatic deacetylation of chitin oligomers, but the diversity is limited by the low number of chitin deacetylases available. We have produced specific chitosan oligomers which are deacetylated at the first two units starting from the non-reducing end by the combined use of two different chitin deacetylases, namely NodB from Rhizobium sp. GRH2 that deacetylates the first unit and COD from Vibrio cholerae that deacetylates the second unit starting from the non-reducing end. Both chitin deacetylases accept the product of each other resulting in production of chitosan oligomers with a novel and defined PA. When extended to further chitin deacetylases, this approach has the potential to yield a large range of novel chitosan oligomers with a fully defined architecture. PMID:25732514

  8. Enzymatic production of defined chitosan oligomers with a specific pattern of acetylation using a combination of chitin oligosaccharide deacetylases

    NASA Astrophysics Data System (ADS)

    Hamer, Stefanie Nicole; Cord-Landwehr, Stefan; Biarnés, Xevi; Planas, Antoni; Waegeman, Hendrik; Moerschbacher, Bruno Maria; Kolkenbrock, Stephan

    2015-03-01

    Chitin and chitosan oligomers have diverse biological activities with potentially valuable applications in fields like medicine, cosmetics, or agriculture. These properties may depend not only on the degrees of polymerization and acetylation, but also on a specific pattern of acetylation (PA) that cannot be controlled when the oligomers are produced by chemical hydrolysis. To determine the influence of the PA on the biological activities, defined chitosan oligomers in sufficient amounts are needed. Chitosan oligomers with specific PA can be produced by enzymatic deacetylation of chitin oligomers, but the diversity is limited by the low number of chitin deacetylases available. We have produced specific chitosan oligomers which are deacetylated at the first two units starting from the non-reducing end by the combined use of two different chitin deacetylases, namely NodB from Rhizobium sp. GRH2 that deacetylates the first unit and COD from Vibrio cholerae that deacetylates the second unit starting from the non-reducing end. Both chitin deacetylases accept the product of each other resulting in production of chitosan oligomers with a novel and defined PA. When extended to further chitin deacetylases, this approach has the potential to yield a large range of novel chitosan oligomers with a fully defined architecture.

  9. Enzymatic production of defined chitosan oligomers with a specific pattern of acetylation using a combination of chitin oligosaccharide deacetylases.

    PubMed

    Hamer, Stefanie Nicole; Cord-Landwehr, Stefan; Biarnés, Xevi; Planas, Antoni; Waegeman, Hendrik; Moerschbacher, Bruno Maria; Kolkenbrock, Stephan

    2015-03-03

    Chitin and chitosan oligomers have diverse biological activities with potentially valuable applications in fields like medicine, cosmetics, or agriculture. These properties may depend not only on the degrees of polymerization and acetylation, but also on a specific pattern of acetylation (PA) that cannot be controlled when the oligomers are produced by chemical hydrolysis. To determine the influence of the PA on the biological activities, defined chitosan oligomers in sufficient amounts are needed. Chitosan oligomers with specific PA can be produced by enzymatic deacetylation of chitin oligomers, but the diversity is limited by the low number of chitin deacetylases available. We have produced specific chitosan oligomers which are deacetylated at the first two units starting from the non-reducing end by the combined use of two different chitin deacetylases, namely NodB from Rhizobium sp. GRH2 that deacetylates the first unit and COD from Vibrio cholerae that deacetylates the second unit starting from the non-reducing end. Both chitin deacetylases accept the product of each other resulting in production of chitosan oligomers with a novel and defined PA. When extended to further chitin deacetylases, this approach has the potential to yield a large range of novel chitosan oligomers with a fully defined architecture.

  10. Production of a low calorie mandarin juice by enzymatic conversion of constituent sugars to oligosaccharides and prevention of insoluble glucan formation.

    PubMed

    Nguyen, Thi Thanh Hanh; Cho, Jae-Young; Seo, Ye-Seul; Woo, Hye-Jin; Kim, Hwa-Kyoung; Kim, Ghahyun J; Jhon, Deok-Young; Kim, Doman

    2015-03-01

    Over 99% of sucrose in mandarin juice (57.1 g/l in original juice to 428.4 g/l in concentrated juice) was enzymatically converted to glucooligosaccharides using 3 U dextransucrase/ml prepared from Leuconostoc mesenteroides at 28 °C. The oligosaccharide synthesis yields were 51 and 47% for the original and the concentrated mandarin juice, respectively. The degree of polymerization of oligosaccharides in the enzyme-modified juice was 2-7. Calories in the original and modified mandarin juice were 433 and 301 kcal/l (30.5% reduction). Compared with the original juice, the enzyme-modified juice showed 82% decrease of insoluble glucan formation by mutansucrase from Streptococcus mutans. A sensory evaluation of the juices revealed that the original and modified mandarin juices had sweetness values of 4.5 and 4.9 and the same values for overall acceptability.

  11. Immobilization of xylanase on poly (ethylene glycol) methyl ether 5000 and its self-extractive bioconversion for the production of xylo-oligosaccharides.

    PubMed

    Li, Xin; Shan, Zongxing; Song, Xiangyang; Ouyang, Jia; Xu, Yong; Yong, Qiang; Yu, Shiyuan

    2014-02-01

    Endo-β-1,4-xylanase derived from Trichoderma reesei was covalently immobilized on poly (ethylene glycol) methyl ether 5000 (mPEG5000), and the resulting immobilized enzyme had a residual activity of 72.4 % with 82.9 % of PEGylated amino groups. Compared with the free enzyme, the immobilized xylanase was stable at pH values in the range of 4.0-6.0 and temperatures in the range of 50-65 °C. A self-extractive bioconversion system composed of immobilized xylanase, mPEG5000, and sodium citrate was used to produce xylo-oligosaccharides and provided a better distribution of the xylo-oligosaccharides than the free enzyme. Furthermore, the immobilized xylanase could be effectively recovered in situ following the hydrolysis reaction.

  12. Dietary Mannan Oligosaccharides: Counteracting the Side Effects of Soybean Meal Oil Inclusion on European Sea Bass (Dicentrarchus labrax) Gut Health and Skin Mucosa Mucus Production?

    PubMed Central

    Torrecillas, Silvia; Montero, Daniel; Caballero, Maria José; Pittman, Karin A.; Custódio, Marco; Campo, Aurora; Sweetman, John; Izquierdo, Marisol

    2015-01-01

    The main objective of this study was to assess the effects of 4 g kg−1 dietary mannan oligosaccharides (MOS) inclusion in soybean oil (SBO)- and fish oil (FO)-based diets on the gut health and skin mucosa mucus production of European sea bass juveniles after 8 weeks of feeding. Dietary MOS, regardless of the oil source, promoted growth. The intestinal somatic index was not affected, however dietary SBO reduced the intestinal fold length, while dietary MOS increased it. The dietary oil source fed produced changes on the posterior intestine fatty acid profiles irrespective of MOS dietary supplementation. SBO down-regulated the gene expression of TCRβ, COX2, IL-1β, TNFα, IL-8, IL-6, IL-10, TGFβ, and Ig and up-regulated MHCII. MOS supplementation up-regulated the expression of MHCI, CD4, COX2, TNFα, and Ig when included in FO-based diets. However, there was a minor up-regulating effect on these genes when MOS was supplemented in the SBO-based diet. Both dietary oil sources and MOS affected mean mucous cell areas within the posterior gut, however the addition of MOS to a SBO diet increased the mucous cell size over the values shown in FO fed fish. Dietary SBO also trends to reduce mucous cell density in the anterior gut relative to FO, suggesting a lower overall mucosal secretion. There are no effects of dietary oil or MOS in the skin mucosal patterns. Complete replacement of FO by SBO, modified the gut fatty acid profile, altered posterior gut-associated immune system (GALT)-related gene expression and gut mucous cells patterns, induced shorter intestinal folds and tended to reduce European sea bass growth. However, when combined with MOS, the harmful effects of SBO appear to be partially balanced by moderating the down-regulation of certain GALT-related genes involved in the functioning of gut mucous barrier and increasing posterior gut mucous cell diffusion rates, thus helping to preserve immune homeostasis. This denotes the importance of a balanced

  13. Dietary Mannan Oligosaccharides: Counteracting the Side Effects of Soybean Meal Oil Inclusion on European Sea Bass (Dicentrarchus labrax) Gut Health and Skin Mucosa Mucus Production?

    PubMed

    Torrecillas, Silvia; Montero, Daniel; Caballero, Maria José; Pittman, Karin A; Custódio, Marco; Campo, Aurora; Sweetman, John; Izquierdo, Marisol

    2015-01-01

    The main objective of this study was to assess the effects of 4 g kg(-1) dietary mannan oligosaccharides (MOS) inclusion in soybean oil (SBO)- and fish oil (FO)-based diets on the gut health and skin mucosa mucus production of European sea bass juveniles after 8 weeks of feeding. Dietary MOS, regardless of the oil source, promoted growth. The intestinal somatic index was not affected, however dietary SBO reduced the intestinal fold length, while dietary MOS increased it. The dietary oil source fed produced changes on the posterior intestine fatty acid profiles irrespective of MOS dietary supplementation. SBO down-regulated the gene expression of TCRβ, COX2, IL-1β, TNFα, IL-8, IL-6, IL-10, TGFβ, and Ig and up-regulated MHCII. MOS supplementation up-regulated the expression of MHCI, CD4, COX2, TNFα, and Ig when included in FO-based diets. However, there was a minor up-regulating effect on these genes when MOS was supplemented in the SBO-based diet. Both dietary oil sources and MOS affected mean mucous cell areas within the posterior gut, however the addition of MOS to a SBO diet increased the mucous cell size over the values shown in FO fed fish. Dietary SBO also trends to reduce mucous cell density in the anterior gut relative to FO, suggesting a lower overall mucosal secretion. There are no effects of dietary oil or MOS in the skin mucosal patterns. Complete replacement of FO by SBO, modified the gut fatty acid profile, altered posterior gut-associated immune system (GALT)-related gene expression and gut mucous cells patterns, induced shorter intestinal folds and tended to reduce European sea bass growth. However, when combined with MOS, the harmful effects of SBO appear to be partially balanced by moderating the down-regulation of certain GALT-related genes involved in the functioning of gut mucous barrier and increasing posterior gut mucous cell diffusion rates, thus helping to preserve immune homeostasis. This denotes the importance of a balanced

  14. A novel thermostable GH5_7 β-mannanase from Bacillus pumilus GBSW19 and its application in manno-oligosaccharides (MOS) production.

    PubMed

    Zang, Haoyu; Xie, Shanshan; Wu, Huijun; Wang, Weiduo; Shao, Xiankun; Wu, Liming; Rajer, Faheem Uddin; Gao, Xuewen

    2015-10-01

    A novel thermostable mannanase from a newly isolated Bacillus pumilus GBSW19 has been identified, expressed, purified and characterized. The enzyme shows a structure comprising a 28 amino acid signal peptide, a glycoside hydrolase family 5 (GH5) catalytic domain and no carbohydrate-binding module. The recombinant mannanase has molecular weight of 45 kDa with an optimal pH around 6.5 and is stable in the range from pH 5-11. Meanwhile, the optimal temperature is around 65 °C, and it retains 50% relative activity at 60 °C for 12h. In addition, the purified enzyme can be activated by several ions and organic solvents and is resistant to detergents. Bpman5 can efficiently convert locus bean gum to mainly M2, M3 and M5, and hydrolyze manno-oligosaccharides with a minimum DP of 3. Further exploration of the optimum condition using HPLC to prepare oligosaccharides from locust bean gum was obtained as 10mg/ml locust bean gum incubated with 10 U/mg enzyme at 50 °C for 24h. By using this enzyme, locust bean gum can be utilized to generate high value-added oligosaccharides with a DP of 2-6. Copyright © 2015 Elsevier Inc. All rights reserved.

  15. Oligosaccharide formation during commercial pear juice processing.

    PubMed

    Willems, Jamie L; Low, Nicholas H

    2016-08-01

    The effect of enzyme treatment and processing on the oligosaccharide profile of commercial pear juice samples was examined by high performance anion exchange chromatography with pulsed amperometric detection and capillary gas chromatography with flame ionization detection. Industrial samples representing the major stages of processing produced with various commercial enzyme preparations were studied. Through the use of commercially available standards and laboratory scale enzymatic hydrolysis of pectin, starch and xyloglucan; galacturonic acid oligomers, glucose oligomers (e.g., maltose and cellotriose) and isoprimeverose were identified as being formed during pear juice production. It was found that the majority of polysaccharide hydrolysis and oligosaccharide formation occurred during enzymatic treatment at the pear mashing stage and that the remaining processing steps had minimal impact on the carbohydrate-based chromatographic profile of pear juice. Also, all commercial enzyme preparations and conditions (time and temperature) studied produced similar carbohydrate-based chromatographic profiles.

  16. Hyper-production and characterization of the ι-carrageenase useful for ι-carrageenan oligosaccharide production from a deep-sea bacterium, Microbulbifer thermotolerans JAMB-A94T, and insight into the unusual catalytic mechanism.

    PubMed

    Hatada, Yuji; Mizuno, Masahiro; Li, Zhijun; Ohta, Yukari

    2011-06-01

    A gene of unknown function from the genome of the agar-degrading deep-sea bacterium Microbulbifer thermotolerans JAMB-A94(T) was functionally identified as a ι-carrageenase gene. This gene, designated as cgiA, is located together with two β-agarase genes, agaA and agaO in a cluster. The cgiA gene product is 569 amino acids and shares 29% identity over 185 amino acids with the ι-carrageenase from Zobellia galactanivorans Dsij DSM 12802. Recombinant, cgiA-encoded ι-carrageenase (55 kDa) was hyper-produced in Bacillus subtilis. The recombinant enzyme shows maximal activity at 50°C, the highest reported optimal temperature for a carrageenase. It cleaved β-1,4 linkages in ι-carrageenan to produce a high ratio of ι-carrageenan tetramer, more than 75% of the total product, and did not cleave the β-1,4 linkages in κ- or λ-carrageenan. Therefore, this enzyme may be useful for industrial production of ι-carrageenan oligosaccharides, which have demonstrated antiviral potential against diverse viruses. Furthermore, we performed site-directed mutagenesis on the gene to identify the catalytic amino acid residues. We demonstrated that a conserved Glu351 was essential for catalysis; however, this enzyme lacked a catalytic Asp residue, which is generally critical for the catalytic activity of most glycoside hydrolases.

  17. Inulin Potential for Enzymatic Obtaining of Prebiotic Oligosaccharides.

    PubMed

    Flores, Adriana C; Morlett, Jesús A; Rodríguez, Raúl

    2016-08-17

    Oligosaccharides have been marketed since the 80s as low-calorie agents and recently have gained interest in the pharmaceutical and food industry as functional sweeteners and prebiotic enriching population of Bifidobacteria. Currently, they have an approximated value of $200 per kg and recently, inulin has been proposed as a feedstock for production of oligosaccharides through selective hydrolysis by action of endoinulinase. High optimum temperature (60°C) and thermostability are two important criteria that determine suitability of this enzyme for industrial applications as well as enzyme cost, a major limiting factor. Significant reduction in cost can be achieved by employing low-value and abundant inulin-rich plants as Jerusalem artichoke, dahlia, yacon, garlic, and onion, among others. In general, the early harvested tubers of these plants contain a greater amount of highly polymerized sugar fractions, which offer more industrial value than late-harvested tubers or those after storage. Also, development of recombinant microorganisms could be useful to reduce the cost of enzyme technology for large-scale production of oligosaccharides. In the case of fungal inulinases, several studies of cloning and modification have been made to achieve greater efficiency. The present paper reviews inulin from vegetable sources as feedstock for oligosaccharides production through the action of inulinases, the impact of polymerization degree of inulin and its availability, and some strategies to increase oligosaccharide production.

  18. Prebiotic oligosaccharides reduce proinflammatory cytokines in intestinal Caco-2 cells via activation of PPARγ and peptidoglycan recognition protein 3.

    PubMed

    Zenhom, Marwa; Hyder, Ayman; de Vrese, Michael; Heller, Knut J; Roeder, Thomas; Schrezenmeir, Jürgen

    2011-05-01

    Prebiotic oligosaccharides modulate the intestinal microbiota and beneficially affect the human body by reducing intestinal inflammation. This immunomodulatory effect was assumed to be bacterial in origin. However, some observations suggest that oligosaccharides may exert an antiinflammatory effect per se. We hypothesized that oligosaccharides affect the intestinal immunity via activation of peptidoglycan recognition protein 3 (PGlyRP3), which reduces the expression of proinflammatory cytokines. Caco-2 cells were treated with the oligosaccharides, α3-sialyllactose, or fructooligosaccharides (Raftilose p95), and the effects of these treatments on PGlyRP3 and PPARγ expression, the release and expression of some proinflammatory cytokines, and NF-κB translocation were tested. Both oligosaccharides had antiinflammatory activity; they significantly reduced IL-12 secretion in Caco-2 cells and gene expression of IL-12p35, IL-8, and TNFα. They also reduced the gene expression and nuclear translocation of NF-κB. Both oligosaccharides dose and time dependently induced the production of PGlyRP3, the silencing of which by transfection of Caco-2 cells with specific small interfering RNA targeting PGlyRP3 abolished the antiinflammatory role of both oligosaccharides. Incubation of Caco-2 cells with both oligosaccharides induced PPARγ. Antagonizing PPARγ by culturing the cells with GW9662 for 24 h inhibited the oligosaccharide-induced PGlyRP3 production and the antiinflammatory effect of the oligosaccharides. We conclude that oligosaccharides may exert an antiinflammatory effect by inducing the nuclear receptor PPARγ, which regulates the antiinflammatory PGlyRP3.

  19. Neutral and Acidic Oligosaccharides Supplementation Does Not Increase the Vaccine Antibody Response in Preterm Infants in a Randomized Clinical Trial

    PubMed Central

    van den Berg, Jolice P.; Westerbeek, Elisabeth A. M.; van der Klis, Fiona R. M.; Berbers, Guy A. M.; Lafeber, Harrie N.; van Elburg, Ruurd M.

    2013-01-01

    Background In preterm infants, a decreased immunological response and lower serological effectiveness are observed after immunizations due to ineffectiveness of both humoral and cellular immune mechanisms. Objective To determine the effect of 80% neutral oligosaccharides [small-chain galacto-oligosaccharides/long-chain fructo-oligosaccharides (scGOS/lcFOS)] in combination with 20% pectin-derived acidic oligosaccharides (pAOS) on antibody concentrations after DTaP-IPV-Hib immunization in preterm infants. Design In this randomized clinical trial, preterm infants with gestational age <32 weeks and/or birth weight <1500 g received enteral supplementation with scGOS/lcFOS/pAOS or placebo (maltodextrin) between days 3 and 30 of life. Blood samples were collected at 5 and 12 months of age. Results In total, 113 infants were included. Baseline and nutritional characteristics were not different in both groups. Geometric mean titers were not different after prebiotic supplementation at 5 months, Ptx (37/44 EU/ml), FHA (78/96 EU/ml), Prn (78/80 EU/ml), Diphtheria (0.40/0.57 IU/ml), Tetanus (0.74/0.99 IU/ml) and Hib (0.35/0.63 µg/ml), and at 12 months Ptx (55/66 EU/ml), FHA (122/119 EU/ml), Prn (116/106 Eu/ml), Diphtheria (0.88/1.11 IU/ml), Tetanus (1.64/1.79 IU/ml) and Hib (2.91/2.55 µg/ml). Conclusions Enteral supplementation of neutral (scGOS/lcFOS) and acidic oligosaccharides (pAOS) does not improve the immunization response in preterm infants. Trial Registration Controlled-Trials.com ISRCTN16211826 ISRCTN16211826 PMID:23951035

  20. Hexose-6-kinases in germinating honey locust cotyledons: substrate specificity of D-fructo-6-kinase.

    PubMed

    Myers, D; Matheson, N K

    1994-11-01

    Extracts of the cotyledons of germinated honey locust (Gleditsia triacanthos) seeds, which contain galactomannan as a reserve polysaccharide in the endosperm, were fractionated by chromatography and the fractions examined for the presence of a specific manno-6-kinase which could phosphorylate the D-mannose released by hydrolysis of galactomannan. One particulate hexokinase (the major hexose-6-kinase fraction) and two soluble hexokinase fractions (the minor portion), as well as a soluble fructo-6-kinase fraction, were initially separated. From chromatography, electrophoresis and kinetic studies, no evidence for a specific manno-kinase was obtained. This and the level and kinetic behaviour of the particulate hexokinase implicated it as the enzyme catalysing the phosphorylation of released D-mannose. The fructo-kinase activity was further separated into three fractions. Kinetic studies on one of these with native and synthetic substrates indicated that the structural requirements for the monosaccharide substrate were a beta-D-anomeric 2-OH in the furanose ring, a 4-OH trans to the D-5-CH2OH and a -CH2OH substituent on C2 (trans to the 5-CH2OH) which could be modified. The orientation of the hydroxyl on C-3 had only a limited effect.

  1. Studies on the asparagine-linked oligosaccharides from cartilage-specific proteoglycan

    SciTech Connect

    Cioffi, L.C.

    1987-01-01

    Chondrocytes synthesize and secrete a cartilage-specific proteoglycan (PG-H) as one of their major products. This proteoglycan has attached to it several types of carbohydrate chains, including chondroitin sulfate, keratan sulfate, O-linked oligosaccharides, and asparagine-linked oligosaccharides. The asparagine-linked oligosaccharides found on PG-H were investigated in these studies. Methodology was developed for the isolation and separation of standard of standard complex and high mannose type oligosaccharides. This included digesting glycoproteins with N-glycanase and separation of the oligosaccharides according to type by concanavalin-A lectin chromatography. The different oligosaccharide types were then analyzed by high pressure liquid chromatography. This methodology was used in the subsequent studies on the PG-H asparagine-linked oligosaccharides. Initially, the asparagine-linked oligosaccharides recovered from the culture medium (CM) and cell-associated (Ma) fractions of PG-H from of tibial chondrocytes were labeled with (/sup 3/H)-mannose and the oligosaccharides were isolated and analyzed.

  2. Optimization of Oligosaccharide Synthesis from Cellobiose by Dextransucrase

    NASA Astrophysics Data System (ADS)

    Kim, Misook; Day, Donal F.

    There is a growing market for oligosaccharides as sweeteners, prebiotics, anticariogenic compounds, and immunostimulating agents in both food and pharmaceutical industries. Interest in novel carbohydrate-based products has grown because of their reduced toxicity and low immune response. Cellobiose is potentially valuable as a nondigestible sugar. The reaction of cellobiose, as an acceptor with a sucrose as a donor, catalyzed by a dextransucrase from Leuconostoc mesenteroides B-512FMCM, produced a series of cellobio-oligosaccharides. This production system was optimized using a Box-Behnken experimental design for 289 mM of sucrose and 250 mM of cellobiose and 54 U of the enzyme at pH 5.2 and 30 °C, to produce maximum yields of oligosaccharide.

  3. Enzymatic synthesis of novel oligosaccharides from N-acetylsucrosamine and melibiose using Aspergillus niger α-galactosidase, and properties of the products.

    PubMed

    Sakaki, Yohei; Tashiro, Mitsuru; Katou, Moe; Sakuma, Chiseko; Hirano, Takako; Hakamata, Wataru; Nishio, Toshiyuki

    2016-09-01

    Two kinds of oligosaccharides, N-acetylraffinosamine (RafNAc) and N-acetylplanteosamine (PlaNAc), were synthesized from N-acetylsucrosamine and melibiose using the transgalactosylation activity of Aspergillus niger α-galactosidase. RafNAc and PlaNAc are novel trisaccharides in which d-glucopyranose residues in raffinose (Raf) and planteose are replaced with N-acetyl-d-glucosamine. These trisaccharides were more stable in acidic solution than Raf. RafNAc was hydrolyzed more rapidly than Raf by α-galactosidase of green coffee bean. In contrast, RafNAc was not hydrolyzed by Saccharomyces cerevisiae invertase, although Raf was hydrolyzed well by this enzyme. These results indicate that the physicochemical properties and steric structure of RafNAc differ considerably from those of Raf.

  4. Novel High-Molecular Weight Fucosylated Milk Oligosaccharides Identified in Dairy Streams

    PubMed Central

    Mehra, Raj; Barile, Daniela; Marotta, Mariarosaria; Lebrilla, Carlito B.; Chu, Caroline; German, J. Bruce

    2014-01-01

    Oligosaccharides are the third largest component in human milk. This abundance is remarkable because oligosaccharides are not digestible by the newborn, and yet they have been conserved and amplified during evolution. In addition to encouraging the growth of a protective microbiota dominated by bifidobacteria, oligosaccharides have anti-infective activity, preventing pathogens from binding to intestinal cells. Although it would be advantageous adding these valuable molecules to infant milk formula, the technologies to reproduce the variety and complexity of human milk oligosaccharides by enzymatic/organic synthesis are not yet mature. Consequently, there is an enormous interest in alternative sources of these valuable oligosaccharides. Recent research has demonstrated that bovine milk and whey permeate also contain oligosaccharides. Thus, a thorough characterization of oligosaccharides in bovine dairy streams is an important step towards fully assessing their specific functionalities. In this study, bovine milk oligosaccharides (BMOs) were concentrated by membrane filtration from a readily available dairy stream called “mother liquor”, and analyzed by high accuracy MALDI FT-ICR mass spectrometry. The combination of HPLC and accurate mass spectrometry allowed the identification of ideal processing conditions leading to the production of Kg amount of BMO enriched powders. Among the BMOs identified, 18 have high-molecular weight and corresponded in size to the most abundant oligosaccharides present in human milk. Notably 6 oligosaccharides contained fucose, a sugar monomer that is highly abundant in human milk, but is rarely observed in bovine milk. This work shows that dairy streams represent a potential source of complex milk oligosaccharides for commercial development of unique dairy ingredients in functional foods that reproduce the benefits of human milk. PMID:24810963

  5. Novel high-molecular weight fucosylated milk oligosaccharides identified in dairy streams.

    PubMed

    Mehra, Raj; Barile, Daniela; Marotta, Mariarosaria; Lebrilla, Carlito B; Chu, Caroline; German, J Bruce

    2014-01-01

    Oligosaccharides are the third largest component in human milk. This abundance is remarkable because oligosaccharides are not digestible by the newborn, and yet they have been conserved and amplified during evolution. In addition to encouraging the growth of a protective microbiota dominated by bifidobacteria, oligosaccharides have anti-infective activity, preventing pathogens from binding to intestinal cells. Although it would be advantageous adding these valuable molecules to infant milk formula, the technologies to reproduce the variety and complexity of human milk oligosaccharides by enzymatic/organic synthesis are not yet mature. Consequently, there is an enormous interest in alternative sources of these valuable oligosaccharides. Recent research has demonstrated that bovine milk and whey permeate also contain oligosaccharides. Thus, a thorough characterization of oligosaccharides in bovine dairy streams is an important step towards fully assessing their specific functionalities. In this study, bovine milk oligosaccharides (BMOs) were concentrated by membrane filtration from a readily available dairy stream called "mother liquor", and analyzed by high accuracy MALDI FT-ICR mass spectrometry. The combination of HPLC and accurate mass spectrometry allowed the identification of ideal processing conditions leading to the production of Kg amount of BMO enriched powders. Among the BMOs identified, 18 have high-molecular weight and corresponded in size to the most abundant oligosaccharides present in human milk. Notably 6 oligosaccharides contained fucose, a sugar monomer that is highly abundant in human milk, but is rarely observed in bovine milk. This work shows that dairy streams represent a potential source of complex milk oligosaccharides for commercial development of unique dairy ingredients in functional foods that reproduce the benefits of human milk.

  6. Passion fruit by-product and fructooligosaccharides stimulate the growth and folate production by starter and probiotic cultures in fermented soymilk.

    PubMed

    Albuquerque, Marcela Albuquerque Cavalcanti; Bedani, Raquel; LeBlanc, Jean Guy; Saad, Susana Marta Isay

    2017-09-05

    Two starter cultures (Streptococcus (St.) thermophilus ST-M6 and TA-40) and five probiotic strains (St. thermophilus TH-4, Lactobacillus (Lb.) acidophilus LA-5, Lb. rhamnosus LGG, Lb. fermentum PCC, and Lb. reuteri RC-14) were used to ferment different soymilk formulations supplemented with passion fruit by-product and/or fructo-oligosaccharides (FOS) with the aim of increasing folate concentrations. Growth and folate production of individual strains were evaluated and the results used to select co-cultures. Both St. thermophilus ST-M6 and TH-4 were the best folate producers and were able to increase the folate content of all soymilk formulations when used alone or in co-culture with lactobacilli strains, especially in the presence of both passion fruit by-product and FOS. Thus, passion fruit by-product and FOS could be used as dietary ingredients to stimulate the folate production by selected bacterial strains during the fermentation of soymilk. It was also shown that vitamin production by microorganisms is strain-dependent and may also be influenced by nutritional and environmental conditions. Copyright © 2017 Elsevier B.V. All rights reserved.

  7. Evaluation of invertase (B-fructo furanosidase) activity in irradiated Mazafaty dates during storage

    NASA Astrophysics Data System (ADS)

    Zare, Z.; Sohrabpour, M.; Fazeli, T. Z.; Kohan, K. G.

    2002-10-01

    Invertase activity of irradiated and non-irradiated Mazafaty dates during four months storage time has been studied. There are large differences in invertase activity in different dates cultivars. The soft and good quality dates usually have higher activity compared to dry or semi-dry varieties. Irradiated dates with doses 1-5 kGy, which could be used for decontamination and disinfestations of dates with a dose rate of 1.87 Gy/s were used. The samples were stored in two temperatures of 5°C and 25°C for four months. The activity of invertase enzyme was analysed at different time intervals. Inactivation study of invertase (B-fructo furanosidase) activity showed that the invertase is sensitive to temperature, storage time and also inactivation of enzyme occurred in dose range of 10-50 kGy.

  8. Oligosaccharides from Sucrose via Glycansucrases

    USDA-ARS?s Scientific Manuscript database

    Glycansucrases are a class of microbial enzymes that polymerize either the fructosyl or the glucosyl moiety of sucrose to give beta-D-fructans or alpha-D-glucans. They are also capable of transferring fructosyl or glucosyl units to acceptor molecules to yield oligosaccharides. Although the glycosy...

  9. Optimization of hydrolysis conditions for the production of glucomanno-oligosaccharides from konjac using β-mannanase by response surface methodology.

    PubMed

    Chen, Junfan; Liu, Desheng; Shi, Bo; Wang, Hai; Cheng, Yongqiang; Zhang, Wenjing

    2013-03-01

    Glucomanno-oligosaccharides (GMO), usually produced from hydrolysis of konjac tubers with a high content of glucomannan, have a positive effect on Bifidobacterium as well as a variety of other physiological activities. Response surface methodology (RSM) was employed to optimize the hydrolysis time, hydrolysis temperature, pH and enzyme to substrate ratio (E/S) to obtain a high GMO yield from konjac tubers. From the signal-factor experiments, it was concluded that the change in the direct reducing sugar (DRS) is consistent with total reducing sugar (TRS) but contrary to the degree of polymerization (DP). DRS was used as an indicator of the content of GMO in the RSM study. The optimum RSM operating conditions were: reaction time of 3.4 h, reaction temperature of 41.0°C, pH of 7.1 and E/S of 0.49. The results suggested that the enzymatic hydrolysis was enhanced by temperature, pH and incubation time. Model validation showed good agreement between experimental results and the predicted responses. Copyright © 2012 Elsevier Ltd. All rights reserved.

  10. Chromatographic Separations of Enantiomers and Underivatized Oligosaccharides

    SciTech Connect

    Liu, Ying

    2004-01-01

    resolution, high column stability, and high sensitivity. In addition, this method showed potential usefulness for the sensitive and quick analysis of hydrolysis products of polysaccharides, and for trace level analysis of individual oligosaccharides or oligosaccharide isomers from biological systems.

  11. Sugar loss and enzyme inhibition due to oligosaccharide accumulation during high solids-loading enzymatic hydrolysis

    SciTech Connect

    Xue, Saisi; Uppugundla, Nirmal; Bowman, Michael J.; Cavalier, David; Da Costa Sousa, Leonardo; Dale, Bruce E.; Balan, Venkatesh

    2015-11-26

    Accumulation of recalcitrant oligosaccharides during high-solids loading enzymatic hydrolysis of cellulosic biomass reduces biofuel yields and increases processing costs for a cellulosic biorefinery. Recalcitrant oligosaccharides in AFEX-pretreated corn stover hydrolysate accumulate to the extent of about 18–25 % of the total soluble sugars in the hydrolysate and 12–18 % of the total polysaccharides in the inlet biomass (untreated), equivalent to a yield loss of about 7–9 kg of monomeric sugars per 100 kg of inlet dry biomass (untreated). These oligosaccharides represent a yield loss and also inhibit commercial hydrolytic enzymes, with both being serious bottlenecks for economical biofuel production from cellulosic biomass. Very little is understood about the nature of these oligomers and why they are recalcitrant to commercial enzymes. This work presents a robust method for separating recalcitrant oligosaccharides from high solid loading hydrolysate in gramme quantities. Composition analysis, recalcitrance study and enzyme inhibition study were performed to understand their chemical nature. Results indicate that, oligosaccharide accumulation occurs during high solid loading enzymatic hydrolysis of corn stover (CS) irrespective of using different pretreated corn stover (dilute acid: DA, ionic liquids: IL, and ammonia fibre expansion: AFEX). The methodology for large-scale separation of recalcitrant oligosaccharides from 25 % solids-loading AFEXcorn stover hydrolysate using charcoal fractionation and size exclusion chromatography is reported for the first time. Oligosaccharides with higher degree of polymerization (DP) were recalcitrant towards commercial enzyme mixtures [Ctec2, Htec2 and Multifect pectinase (MP)] compared to lower DP oligosaccharides. Enzyme inhibition studies using processed substrates (Avicel and xylan) showed that low DP oligosaccharides also inhibit commercial enzymes. Addition of monomeric sugars to oligosaccharides increases the

  12. Sugar loss and enzyme inhibition due to oligosaccharide accumulation during high solids-loading enzymatic hydrolysis

    DOE PAGES

    Xue, Saisi; Uppugundla, Nirmal; Bowman, Michael J.; ...

    2015-11-26

    Accumulation of recalcitrant oligosaccharides during high-solids loading enzymatic hydrolysis of cellulosic biomass reduces biofuel yields and increases processing costs for a cellulosic biorefinery. Recalcitrant oligosaccharides in AFEX-pretreated corn stover hydrolysate accumulate to the extent of about 18–25 % of the total soluble sugars in the hydrolysate and 12–18 % of the total polysaccharides in the inlet biomass (untreated), equivalent to a yield loss of about 7–9 kg of monomeric sugars per 100 kg of inlet dry biomass (untreated). These oligosaccharides represent a yield loss and also inhibit commercial hydrolytic enzymes, with both being serious bottlenecks for economical biofuel production frommore » cellulosic biomass. Very little is understood about the nature of these oligomers and why they are recalcitrant to commercial enzymes. This work presents a robust method for separating recalcitrant oligosaccharides from high solid loading hydrolysate in gramme quantities. Composition analysis, recalcitrance study and enzyme inhibition study were performed to understand their chemical nature. Results indicate that, oligosaccharide accumulation occurs during high solid loading enzymatic hydrolysis of corn stover (CS) irrespective of using different pretreated corn stover (dilute acid: DA, ionic liquids: IL, and ammonia fibre expansion: AFEX). The methodology for large-scale separation of recalcitrant oligosaccharides from 25 % solids-loading AFEXcorn stover hydrolysate using charcoal fractionation and size exclusion chromatography is reported for the first time. Oligosaccharides with higher degree of polymerization (DP) were recalcitrant towards commercial enzyme mixtures [Ctec2, Htec2 and Multifect pectinase (MP)] compared to lower DP oligosaccharides. Enzyme inhibition studies using processed substrates (Avicel and xylan) showed that low DP oligosaccharides also inhibit commercial enzymes. Addition of monomeric sugars to oligosaccharides increases the

  13. Intestinal infections and prebiotics: the roles of oligosaccharides in promoting health

    USDA-ARS?s Scientific Manuscript database

    Prebiotic oligosaccharides exert activity against pathogens partly by stimulating the growth and/or activity of commensal bacteria that provide health benefits (lower pH, bacteriocin production, immune system modulation, competitive exclusion). This review describes alternative mechanisms of action...

  14. The Human Milk Metabolome Reveals Diverse Oligosaccharide Profiles123

    PubMed Central

    Smilowitz, Jennifer T.; O’Sullivan, Aifric; Barile, Daniela; German, J. Bruce; Lönnerdal, Bo; Slupsky, Carolyn M.

    2013-01-01

    Breast milk delivers nutrition and protection to the developing infant. There has been considerable research on the high-molecular-weight milk components; however, low-molecular-weight metabolites have received less attention. To determine the effect of maternal phenotype and diet on the human milk metabolome, milk collected at day 90 postpartum from 52 healthy women was analyzed by using proton nuclear magnetic resonance spectroscopy. Sixty-five milk metabolites were quantified (mono-, di-, and oligosaccharides; amino acids and derivatives; energy metabolites; fatty acids and associated metabolites; vitamins, nucleotides, and derivatives; and others). The biological variation, represented as the percentage CV of each metabolite, varied widely (4–120%), with several metabolites having low variation (<20%), including lactose, urea, glutamate, myo-inositol, and creatinine. Principal components analysis identified 2 clear groups of participants who were differentiable on the basis of milk oligosaccharide concentration and who were classified as secretors or nonsecretors of fucosyltransferase 2 (FUT2) gene products according to the concentration of 2′-fucosyllactose, lactodifucotetraose, and lacto-N-fucopentaose I. Exploration of the interrelations between the milk sugars by using Spearman rank correlations revealed significant positive and negative associations, including positive correlations between fucose and products of the FUT2 gene and negative correlations between fucose and products of the fucosyltransferase 3 (FUT3) gene. The total concentration of milk oligosaccharides was conserved among participants (%CV = 18%), suggesting tight regulation of total oligosaccharide production; however, concentrations of specific oligosaccharides varied widely between participants (%CV = 30.4–84.3%). The variability in certain milk metabolites suggests possible roles in infant or infant gut microbial development. This trial was registered at clinicaltrials.gov as NCT

  15. The human milk metabolome reveals diverse oligosaccharide profiles.

    PubMed

    Smilowitz, Jennifer T; O'Sullivan, Aifric; Barile, Daniela; German, J Bruce; Lönnerdal, Bo; Slupsky, Carolyn M

    2013-11-01

    Breast milk delivers nutrition and protection to the developing infant. There has been considerable research on the high-molecular-weight milk components; however, low-molecular-weight metabolites have received less attention. To determine the effect of maternal phenotype and diet on the human milk metabolome, milk collected at day 90 postpartum from 52 healthy women was analyzed by using proton nuclear magnetic resonance spectroscopy. Sixty-five milk metabolites were quantified (mono-, di-, and oligosaccharides; amino acids and derivatives; energy metabolites; fatty acids and associated metabolites; vitamins, nucleotides, and derivatives; and others). The biological variation, represented as the percentage CV of each metabolite, varied widely (4-120%), with several metabolites having low variation (<20%), including lactose, urea, glutamate, myo-inositol, and creatinine. Principal components analysis identified 2 clear groups of participants who were differentiable on the basis of milk oligosaccharide concentration and who were classified as secretors or nonsecretors of fucosyltransferase 2 (FUT2) gene products according to the concentration of 2'-fucosyllactose, lactodifucotetraose, and lacto-N-fucopentaose I. Exploration of the interrelations between the milk sugars by using Spearman rank correlations revealed significant positive and negative associations, including positive correlations between fucose and products of the FUT2 gene and negative correlations between fucose and products of the fucosyltransferase 3 (FUT3) gene. The total concentration of milk oligosaccharides was conserved among participants (%CV = 18%), suggesting tight regulation of total oligosaccharide production; however, concentrations of specific oligosaccharides varied widely between participants (%CV = 30.4-84.3%). The variability in certain milk metabolites suggests possible roles in infant or infant gut microbial development. This trial was registered at clinicaltrials.gov as NCT01817127.

  16. Automated sample preparation facilitated by PhyNexus MEA purification system for oligosaccharide mapping of glycoproteins.

    PubMed

    Prater, Bradley D; Anumula, Kalyan R; Hutchins, Jeff T

    2007-10-15

    A reproducible high-throughput sample cleanup method for fluorescent oligosaccharide mapping of glycoproteins is described. Oligosaccharides are released from glycoproteins using PNGase F and labeled with 2-aminobenzoic acid (anthranilic acid, AA). A PhyNexus MEA system was adapted for automated isolation of the fluorescently labeled oligosaccharides from the reaction mixture prior to mapping by HPLC. The oligosaccharide purification uses a normal-phase polyamide resin (DPA-6S) in custom-made pipette tips. The resin volume, wash, and elution steps involved were optimized to obtain high recovery of oligosaccharides with the least amount of contaminating free fluorescent dye in the shortest amount of time. The automated protocol for sample cleanup eliminated all manual manipulations with a recycle time of 23 min. We have reduced the amount of excess AA by 150-fold, allowing quantitative oligosaccharide mapping from as little as 500 ng digested recombinant immunoglobulin G (rIgG). This low sample requirement allows early selection of a cell line with desired characteristics (e.g., oligosaccharide profile and high specific productivity) for the production of glycoprotein drugs. In addition, the use of Tecan or another robotic platform in conjunction with this method should allow the cleanup of 96 samples in 23 min, a significant decrease in the amount of time currently required to process such a large number of samples.

  17. Inhibition of high-molecular-weight-(1-->3)-beta-D-glucan-dependent activation of a limulus coagulation factor G by laminaran oligosaccharides and curdlan degradation products.

    PubMed

    Tanaka, S; Aketagawa, J; Takahashi, S; Shibata, Y; Tsumuraya, Y; Hashimoto, Y

    1993-05-21

    Extensive surveys for the effects of various beta-D-glucans on the coagulation cascade in horseshoe crab amebocyte lysates showed that low-mol-wt-(1-->3)-beta-D-glucans and laminaran oligosaccharides inhibit the activation of a limulus coagulation factor G by high-mol-wt-(1-->3)-beta-D-glucans. The inhibitory properties are exclusively dependent upon their number-average mol wt (Mn) in a range of 342-58,100, which correspond to a degree of polymerization (dp) range of 2-359. The most effective is a laminaran dextrin of Mn 5800 (dp of 35-36), which causes 50% inhibition of factor G activation at a concentration of 3.16 ng/mL. The inhibition of the activation of factor G proportional to the concentration of the inhibitor, and the adsorption of factor G by inhibitory beta-D-glucan-conjugated cellulose suggested a high affinity of the inhibitory saccharides for the activator-recognition site of factor G. Branched (1-->6), (1-->3)-beta-D-glucans, laminarans, mixed linkage (1-->3), (1-->4)-beta-D-glucans, and partially substituted curdlan and laminaran were found to be inhibitory, possibly owing to clusters of consecutive (1-->3)-beta-D-glucopyranosyl residues as intrachain units. The inhibition appears to be related to the inability of the inhibitory (1-->3)-beta-D-glucans to form ordered conformations and to their tendency to take a random-coil structure in aqueous solution.

  18. Prebiotic effects of diet supplemented with the cultivated red seaweed Chondrus crispus or with fructo-oligo-saccharide on host immunity, colonic microbiota and gut microbial metabolites.

    PubMed

    Liu, Jinghua; Kandasamy, Saveetha; Zhang, Junzeng; Kirby, Christopher W; Karakach, Tobias; Hafting, Jeff; Critchley, Alan T; Evans, Franklin; Prithiviraj, Balakrishnan

    2015-08-14

    Gastrointestinal microbial communities are diverse and are composed of both beneficial and pathogenic groups. Prebiotics, such as digestion-resistant fibers, influence the composition of gut microbiota, and can contribute to the improvement of host health. The red seaweed Chondrus crispus is rich in dietary fiber and oligosaccharides, however its prebiotic potential has not been studied to date. Prebiotic effects were investigated with weaning rats fed a cultivated C. crispus-supplemented diet. Comparison standards included a fructo-oligo-saccharide (FOS) diet and a basal diet. The colonic microbiome was profiled with a 16S rRNA sequencing-based Phylochip array. Concentrations of short chain fatty acids (SCFAs) in the feacal samples were determined by gas chromatography with a flame ionization detector (GC-FID) analysis. Immunoglobulin levels in the blood plasma were analyzed with an enzyme-linked immunosorbent assay (ELISA). Histo-morphological parameters of the proximal colon tissue were characterized by hematoxylin and eosin (H&E) staining. Phylochip array analysis indicated differing microbiome composition among the diet-supplemented and the control groups, with the C. crispus group (2.5% supplementation) showing larger separation from the control than other treatment groups. In the 2.5% C. crispus group, the population of beneficial bacteria such as Bifidobacterium breve increased (4.9-fold, p=0.001), and the abundance of pathogenic species such as Clostridium septicum and Streptococcus pneumonia decreased. Higher concentrations of short chain fatty acids (i.e., gut microbial metabolites), including acetic, propionic and butyric acids, were found in faecal samples of the C. crispus-fed rats. Furthermore, both C. crispus and FOS supplemented rats showed significant improvements in proximal colon histo-morphology. Higher faecal moisture was noted in the 2.5% C. crispus group, and elevated plasma immunoglobulin (IgA and IgG) levels were observed in the 0.5% C

  19. Xylo- and cello-oligosaccharide oxidation by gluco-oligosaccharide oxidase from Sarocladium strictum and variants with reduced substrate inhibition

    PubMed Central

    2013-01-01

    Background The oxidation of carbohydrates from lignocellulose can facilitate the synthesis of new biopolymers and biochemicals, and also reduce sugar metabolism by lignocellulolytic microorganisms, reserving aldonates for fermentation to biofuels. Although oxidoreductases that oxidize cellulosic hydrolysates have been well characterized, none have been reported to oxidize substituted or branched xylo-oligosaccharides. Moreover, this is the first report that identifies amino acid substitutions leading to GOOX variants with reduced substrate inhibition. Results The recombinant wild type gluco-oligosaccharide oxidase (GOOX) from the fungus Sarocladium strictum, along with variants that were generated by site-directed mutagenesis, retained the FAD cofactor, and showed high activity on cello-oligosaccharide and xylo-oligosaccharides, including substituted and branched xylo-oligosaccharides. Mass spectrometric analyses confirmed that GOOX introduces one oxygen atom to oxidized products, and 1H NMR and tandem mass spectrometry analysis confirmed that oxidation was restricted to the anomeric carbon. The A38V mutation, which is close to a predicted divalent ion-binding site in the FAD-binding domain of GOOX but 30 Å away from the active site, significantly increased the kcat and catalytic efficiency of the enzyme on all oligosaccharides. Eight amino acid substitutions were separately introduced to the substrate-binding domain of GOOX-VN (at positions Y72, E247, W351, Q353 and Q384). In all cases, the Km of the enzyme variant was higher than that of GOOX, supporting the role of corresponding residues in substrate binding. Most notably, W351A increased Km values by up to two orders of magnitude while also increasing kcat up to 3-fold on cello- and xylo-oligosaccharides and showing no substrate inhibition. Conclusions This study provides further evidence that S. strictum GOOX has broader substrate specificity than the enzyme name implies, and that substrate inhibition can be

  20. Plant oligosaccharides - outsiders among elicitors?

    PubMed

    Larskaya, I A; Gorshkova, T A

    2015-07-01

    This review substantiates the need to study the plant oligoglycome. The available information on oligosaccharins - physiologically active fragments of plant cell wall polysaccharides - is summarized. The diversity of such compounds in chemical composition, origin, and proved biological activity is highlighted. At the same time, plant oligosaccharides can be considered as outsiders among elicitors of various natures in research intensity of recent decades. This review discusses the reasons for such attitude towards these regulators, which are largely connected with difficulties in isolation and identification. Together with that, approaches are suggested whose potentials can be used to study oligosaccharins. The topics of oligosaccharide metabolism in plants, including the ways of formation, transport, and inactivation are presented, together with data on biological activity and interaction with plant hormones. The current viewpoints on the mode of oligosaccharin action - perception, signal transduction, and possible "targets" - are considered. The potential uses of such compounds in medicine, food industry, agriculture, and biotechnology are discussed.

  1. Tyrosine derivatization and preparative purification of the sialyl and asialy-N-linked oligosaccharides from porcine fibrinogen.

    PubMed

    Da Silva, M L; Tamura, T; McBroom, T; Rice, K G

    1994-07-01

    The N-linked oligosaccharides from porcine fibrinogen were purified following their release from glycopeptides using N-glycosidase F. In separate experiments, both sialyl and asialyl oligosaccharides were prepared from 5 g of fibrinogen. The reducing oligosaccharides were reacted with ammonium bicarbonate to form/oligosaccharide-glycosylamines and then derivatized with tert-butoxycarbonyl-L-tyrosine N-hydroxysuccinimidyl ester. Tyrosinamide--oligosaccharides were purified first by gel filtration chromatography and then by reverse-phase HPLC and the products were characterized by proton NMR and fast atom bombardment-MS. Porcine fibrinogen was found to have predominantly a single asialyl biantennary oligosaccharide containing a fucose linked alpha 1-6 to GlcNAc 1. The oligosaccharide possesses two sialylation patterns with a major form (70%) having a single N-acetyl neuraminic acid (NeuAc) residue linked alpha 2-6 to galactose on only one antenna and a minor form (30%) possessing two NeuAc residues linked alpha 2-6 to both terminal galactose residues. In addition to developing an isolation procedure and establishing the structures of porcine fibrinogen oligosaccharides, this study improves on the tyrosine derivatization technique as a general approach to isolate structurally diverse N-linked oligosaccharides from glycoproteins.

  2. Bovine Milk as a Source of Functional Oligosaccharides for Improving Human Health12

    PubMed Central

    Zivkovic, Angela M.; Barile, Daniela

    2011-01-01

    Human milk oligosaccharides are complex sugars that function as selective growth substrates for specific beneficial bacteria in the gastrointestinal system. Bovine milk is a potentially excellent source of commercially viable analogs of these unique molecules. However, bovine milk has a much lower concentration of these oligosaccharides than human milk, and the majority of the molecules are simpler in structure than those found in human milk. Specific structural characteristics of milk-derived oligosaccharides are crucial to their ability to selectively enrich beneficial bacteria while inhibiting or being less than ideal substrates for undesirable and pathogenic bacteria. Thus, if bovine milk products are to provide human milk–like benefits, it is important to identify specific dairy streams that can be processed commercially and cost-effectively and that can yield specific oligosaccharide compositions that will be beneficial as new food ingredients or supplements to improve human health. Whey streams have the potential to be commercially viable sources of complex oligosaccharides that have the structural resemblance and diversity of the bioactive oligosaccharides in human milk. With further refinements to dairy stream processing techniques and functional testing to identify streams that are particularly suitable for enriching beneficial intestinal bacteria, the future of oligosaccharides isolated from dairy streams as a food category with substantiated health claims is promising. PMID:22332060

  3. Synthesis and characterization of isomaltulose-derived oligosaccharides produced by transglucosylation reaction of Leuconostoc mesenteroides dextransucrase.

    PubMed

    Barea-Alvarez, Montserrat; Benito, Maria Teresa; Olano, Agustín; Jimeno, Maria Luisa; Moreno, F Javier

    2014-09-17

    This paper reports the efficient enzymatic synthesis of a homologous series of isomaltulose-derived oligosaccharides with degrees of polymerization ranging from 3 to 9 through the transglucosylation reaction using a dextransucrase from Leuconostoc mesenteroides B-512F. The total oligosaccharide yield obtained under optimal conditions was 41-42% (in weight with respect to the initial amount of isomaltulose) after 24-48 h of reaction. Nuclear magnetic resonance (NMR) structural characterization indicated that dextransucrase specifically transferred glucose moieties of sucrose to the C-6 of the nonreducing glucose residue of isomaltulose. Likewise, monitoring the progression of the content of each individual oligosaccharide indicated that oligosaccharide acceptor products of low molecular weight acted in turn as acceptors for further transglucosylation to yield oligosaccharides of a higher degree of polymerization. The produced isomaltulose-derived oligosaccharides can be considered as isomalto-oligosaccharides (IMOs) because they are linked by only α-(1→6) bonds. In addition, having isomaltulose as the core structure, these IMO-like structures could possess appealing bioactive properties that could find potential applications as functional food ingredients.

  4. Bovine milk as a source of functional oligosaccharides for improving human health.

    PubMed

    Zivkovic, Angela M; Barile, Daniela

    2011-05-01

    Human milk oligosaccharides are complex sugars that function as selective growth substrates for specific beneficial bacteria in the gastrointestinal system. Bovine milk is a potentially excellent source of commercially viable analogs of these unique molecules. However, bovine milk has a much lower concentration of these oligosaccharides than human milk, and the majority of the molecules are simpler in structure than those found in human milk. Specific structural characteristics of milk-derived oligosaccharides are crucial to their ability to selectively enrich beneficial bacteria while inhibiting or being less than ideal substrates for undesirable and pathogenic bacteria. Thus, if bovine milk products are to provide human milk-like benefits, it is important to identify specific dairy streams that can be processed commercially and cost-effectively and that can yield specific oligosaccharide compositions that will be beneficial as new food ingredients or supplements to improve human health. Whey streams have the potential to be commercially viable sources of complex oligosaccharides that have the structural resemblance and diversity of the bioactive oligosaccharides in human milk. With further refinements to dairy stream processing techniques and functional testing to identify streams that are particularly suitable for enriching beneficial intestinal bacteria, the future of oligosaccharides isolated from dairy streams as a food category with substantiated health claims is promising.

  5. Analysis and role of oligosaccharides in milk.

    PubMed

    Ruhaak, L Renee; Lebrilla, Carlito B

    2012-08-01

    Milk is an important fluid in glycobiology because it contains a number of short carbohydrate chains either free or as glycoconjugates. These compounds as a class are the most abundant component and benefit the infant by developing and maintaining the infant's gut flora. New and emerging methods for oligosaccharide analysis have been developed to study milk. These methods allow for the rapid profiling of oligosaccharide mixtures with quantitation. With these tools, the role of oligosaccharide in milk is being understood. They further point to how oligosaccharide analysis can be performed, which until now has been very difficult and have lagged significantly those of other biopolymers.

  6. Identification and characterisation of water and alkali soluble oligosaccharides from hazelnut skin (Corylus avellana L.).

    PubMed

    Montella, Rosa; Coïsson, Jean Daniel; Travaglia, Fabiano; Locatelli, Monica; Bordiga, Matteo; Meyrand, Mickael; Barile, Daniela; Arlorio, Marco

    2013-10-15

    Hazelnut skins are a good example of agricultural by-product with the potential to become a valuable source of functional ingredients. In this work, the fibre from hazelnut skins was extracted by using water and alkali solution and characterised by a suite of analytical tools (MALDI-FTICR, nano LC-Chip-Q-ToF and gas chromatography). Over thirty complex free oligosaccharides, composed mainly of galacturonic acid and N-acetylgalactosamine, were characterised for the first time in the present study. Their concentration ranged between 16 and 34mg per g of extract. The oligosaccharides isolated from this agricultural by-product are mainly hexose oligosaccharides (potentially galacto-oligosaccharides,) and xyloglucans. The identified composition could justify the bioactive activity of the extracts, namely prebiotic activity, previously demonstrated.

  7. Production of the Functional Trisaccharide 1-Kestose from Cane Sugar Molasses Using Aspergillus japonicus β-Fructofuranosidase.

    PubMed

    Hirabayashi, Katsuki; Kondo, Nobuhiro; Toyota, Hiroshi; Hayashi, Sachio

    2017-01-01

    We report the production of the functional trisaccharide 1-kestose, O-β-D-fructofuranosyl-(2→1)-β-D-fructofuranosyl α-D-glucopyranoside, by β-fructofuranosidase from Aspergillus japonicus using sugar cane molasses as substrate. Sucrose in cane sugar molasses acted as a fructosyl donor and acceptor for the enzyme. The tetrasaccharide nystose, O-β-D-fructofuranosyl-(2→1)-β-D-fructofuranosyl-(2→1)-β-D-fructofuranosyl α-D-glucopyranoside, was produced from 1-kestose. Cane sugar molasses mixed with water provided a better substrate solution for β-fructofuranosidase compared to undiluted molasses due to the high concentration of product inhibitors such as glucose and fructose in molasses. The maximum concentration of 1-kestose obtained was 84.9 mg/ml and the maximum production efficiency was 32.3% after 24 h reaction at 40 °C. The maximum efficiency of combined fructo-oligosaccharide (1-kestose and nystose) production was 40.6%. 1-Kestose was therefore produced via a fructosyl-transfer reaction catalyzed by β-fructofuranosidase from A. japonicus.

  8. Analysis of Prebiotic Oligosaccharides

    NASA Astrophysics Data System (ADS)

    Sanz, M. L.; Ruiz-Matute, A. I.; Corzo, N.; Martínez-Castro, I.

    Carbohydrates and more specifically prebiotics, are complex mixtures of isomers with different degrees of polymerization (DP), monosaccharide units and/or glycosidic linkages. Many efforts are focused on the search for new products and the determination of their biological activity. However, the study of their chemical structure is fundamental to both acquire a basic knowledge of the carbohydrate and to increase the understanding of the mechanisms for their metabolic effect.

  9. The transfer of mannose to dolichol diphosphate oligosaccharides in pig liver endoplasmic reticulum.

    PubMed

    Oliver, G J; Hemming, F W

    1975-11-01

    The transfer, catalysed by pig liver microsomal preparations, of mannose, from GDP-mannose, to lipid-linked oligosaccharides and the properties of the products are described. Solubility, hydrolytic and chromatographic data suggest that they are dolichol diphosphate derivatives. The presence of two N-acetyl groups in at least part of the heterogenous oligosaccharide portion was tentatively deduced. Reduction with borohydride of the oligosaccharide showed that the newly added mannose residues were not at its reducing end. Periodate oxidation suggested that 60% of these were at the non-reducing terminus and that 40% were positioned internally. T.l.c. showed the presence of seven oligosaccharide fractions with chromatographic mobilities corresponding to glucose oligomers with 7-13 residues. The molar proportions of the oligosaccharide fractions in the mixture were determined by borotritiide reduction and the number of mannose residues added to each oligosaccharide fraction during the incubation was calculated. Two of the oligosaccharide fractions had received on average one, or slightly more than one, mannose residue per chain during the incubation; four of the other fractions were each shown to be a mixture, 20-25% of which had received one mannose residue during the incubation and 75-80% of which had not been mannosylated during the incubation. This supported other evidence for the presence of endogenous lipid-linked oligosaccharides in the microsomal preparation which had been formed before the incubation in vitro. Evidence for the possibility of two pools of dolichol monophosphate mannose, one being more closely associated with mannosyl transfer to dolichol diphosphate oligosaccharides than the other, is also discussed.

  10. A combined metabolomic and phylogenetic study reveals putatively prebiotic effects of high molecular weight arabino-oligosaccharides when assessed by in vitro fermentation in bacterial communities derived from humans.

    PubMed

    Sulek, Karolina; Vigsnaes, Louise Kristine; Schmidt, Line Rieck; Holck, Jesper; Frandsen, Henrik Lauritz; Smedsgaard, Jørn; Skov, Thomas Hjort; Meyer, Anne S; Licht, Tine Rask

    2014-08-01

    Prebiotic oligosaccharides are defined by their selective stimulation of growth and/or activity of bacteria in the digestive system in ways claimed to be beneficial for health. However, apart from the short chain fatty acids, little is known about bacterial metabolites created by fermentation of prebiotics, and the significance of the size of the oligosaccharides remains largely unstudied. By in vitro fermentations in human fecal microbial communities (derived from six different individuals), we studied the effects of high-mass (HA, >1 kDa), low-mass (LA, <1 kDa) and mixed (BA) sugar beet arabino-oligosaccharides (AOS) as carbohydrate sources. Fructo-oligosaccharides (FOS) were included as reference. The changes in bacterial communities and the metabolites produced in response to incubation with the different carbohydrates were analyzed by quantitative PCR (qPCR) and Liquid Chromatography-Mass Spectrometry (LC-MS), respectively. All tested carbohydrate sources resulted in a significant increase of Bifidobacterium spp. between 1.79 fold (HA) and 1.64 fold (FOS) in the microbial populations after fermentation, and LC-MS analysis suggested that the bifidobacteria contributed to decomposition of the arabino-oligosaccharide structures, most pronounced in the HA fraction, resulting in release of the essential amino acid phenylalanine. Abundance of Lactobacillus spp. correlated with the presence of a compound, most likely a flavonoid, indicating that lactobacilli contribute to release of such health-promoting substances from plant structures. Additionally, the combination of qPCR and LC-MS revealed a number of other putative interactions between intestinal microbes and the oligosaccharides, which contributes to the understanding of the mechanisms behind prebiotic impact on human health.

  11. Synthesis and Fermentation Properties of Novel Galacto-Oligosaccharides by β-Galactosidases from Bifidobacterium Species

    PubMed Central

    Rabiu, Bodun A.; Jay, Andrew J.; Gibson, Glenn R.; Rastall, Robert A.

    2001-01-01

    β-Galactosidase enzymes were extracted from pure cultures of Bifidobacterium angulatum, B. bifidum BB-12, B. adolescentis ANB-7, B. infantis DSM-20088, and B. pseudolongum DSM-20099 and used in glycosyl transfer reactions to synthesize oligosaccharides from lactose. At a lactose concentration of 30% (wt/wt) oligosaccharide yields of 24.7 to 47.6% occurred within 7 h. Examination of the products by thin-layer chromatography and methylation analysis revealed distinct product derived spectra from each enzyme. These were found to be different to that of Oligomate 55, a commercial prebiotic galacto-oligosaccharide. Fermentation testing of the oligosaccharides showed an increase in growth rate, compared to Oligomate 55, with products derived from B. angulatum, B. bifidum, B. infantis, and B. pseudolongum. However B. adolescentis had a lower growth rates on its oligosaccharide compared with Oligomate 55. Mixed culture testing of the B. bifidum BS-4 oligosaccharide showed that the overall prebiotic effect was equivalent to that of Oligomate 55. PMID:11375159

  12. Galacto-Oligosaccharide Prebiotics

    NASA Astrophysics Data System (ADS)

    Tzortzis, George; Vulevic, Jelena

    The wide recognition of bifidobacteria as health promoting bacteria (Boesten and de Vos, 2008) has attracted a lot of interest in identifying substances that can selectively promote their growth. Many studies using conventional culture and molecular techniques for bacterial identification have shown that breast-fed infants are characterized by an intestinal microbiota that is dominated by bifidobacteria (Benno et al., 1984), which is different from that of infants fed on cow's milk in that their microbiotas are characterized by lower counts of bifidobacteria, with greater numbers of more potentially harmful organisms such as clostridia and enterococci (Lunderquist et al., 1985). As a result of this difference in the microbiota composition, higher levels of ammonia, amines and phenols and other potentially harmful substances have also been found in infants fed cow's milk products (Lunderquist et al., 1985).

  13. [Molecular structure and fractal analysis of oligosaccharide].

    PubMed

    Liu, Wen-long; Wang, Lu-man; He, Dong-qi; Zhang, Tian-lan; Gou, Bao-di; Li, Qing

    2014-10-18

    To propose a calculation method of oligosaccharides' fractal dimension, and to provide a new approach to studying the drug molecular design and activity. By using the principle of energy optimization and computer simulation technology, the steady structures of oligosaccharides were found, and an effective way of oligosaccharides fractal dimension's calculation was further established by applying the theory of box dimension to the chemical compounds. By using the proposed method, 22 oligosaccharides' fractal dimensions were calculated, with the mean 1.518 8 ± 0.107 2; in addition, the fractal dimensions of the two activity multivalent oligosaccharides which were confirmed by experiments, An-2 and Gu-4, were about 1.478 8 and 1.516 0 respectively, while C-type lectin-like receptor Dectin-1's fractal dimension was about 1.541 2. The experimental and computational results were expected to help to find a class of glycoside drugs whose target receptor was Dectin-1. Fractal dimension, differing from other known macro parameters, is a useful tool to characterize the compound molecules' microscopic structure and function, which may play an important role in the molecular design and biological activity study. In the process of oligosaccharides drug screening, the fractal dimension of receptor and designed oligosaccharides or glycoclusters can be calculated respectively. The oligosaccharides with fractal dimension close to that of target receptor should then take priority compared with others, to get the drug molecules with latent activity.

  14. Effect of a Semi-Purified Oligosaccharide-Enriched Fraction from Caprine Milk on Barrier Integrity and Mucin Production of Co-Culture Models of the Small and Large Intestinal Epithelium

    PubMed Central

    Barnett, Alicia M.; Roy, Nicole C.; McNabb, Warren C.; Cookson, Adrian L.

    2016-01-01

    Caprine milk contains the highest amount of oligosaccharides among domestic animals, which are structurally similar to human milk oligosaccharides (HMOs). This suggests caprine milk oligosaccharides may offer similar protective and developmental effects to that of HMOs. However, to date, studies using oligosaccharides from caprine milk have been limited. Thus, this study aimed to examine the impact of a caprine milk oligosaccharide-enriched fraction (CMOF) on barrier function of epithelial cell co-cultures of absorptive enterocytes (Caco-2 cells) and mucus-secreting goblet cells (HT29-MTX cells), that more closely simulate the cell proportions found in the small (90:10) and large intestine (75:25). Treatment of epithelial co-cultures with 0.4, 1.0, 2.0 and 4.0 mg/mL of CMOF was shown to have no effect on metabolic activity but did enhance cell epithelial barrier integrity as measured by trans-epithelial electrical resistance (TEER), in a dose-dependent manner. The CMOF at the maximum concentration tested (4.0 mg/mL) enhanced TEER, mucin gene expression and mucin protein abundance of epithelial co-cultures, all of which are essential components of intestinal barrier function. PMID:27164134

  15. Energy-Dependent Electron Activated Dissociation of Metal-Adducted Permethylated Oligosaccharides

    PubMed Central

    Yu, Xiang; Huang, Yiqun; Lin, Cheng; Costello, Catherine E.

    2013-01-01

    The effects of varying the electron energy and cationizing agents on electron activated dissociation (ExD) of metal-adducted oligosaccharides were explored, using permethylated maltoheptaose as the model system. Across the examined range of electron energy, the metal-adducted oligosaccharide exhibited several fragmentation processes, including electron capture dissociation (ECD) at low energies, hot-ECD at intermediate energies, and electronic excitation dissociation (EED) at high energies. The dissociation threshold depended on the metal charge carrier(s), whereas the types and sequence spans of product ions were influenced by the metal-oligosaccharide binding pattern. Theoretical modeling contributed insight into the metal-dependent behavior of carbohydrates during low-energy ECD. When ExD was applied to a permethylated high mannose N-linked glycan, EED provided more structural information than either collision-induced dissociation (CID) or low-energy ECD, thus demonstrating its potential for oligosaccharide linkage analysis. PMID:22881449

  16. Development of N- and O-linked oligosaccharide engineered Saccharomyces cerevisiae strain.

    PubMed

    Abe, Hiroko; Tomimoto, Kazuya; Fujita, Yasuko; Iwaki, Tomoko; Chiba, Yasunori; Nakayama, Ken-Ichi; Nakajima, Yoshihiro

    2016-11-01

    Yeast cells have been engineered for the production of glycoproteins as biopharmaceuticals with humanized N-linked oligosaccharides. The suppression of yeast-specific O-mannosylation is important to reduce immune response and to improve heterologous protein productivity in the production of biopharmaceuticals. However, so far, there are few reports of the engineering of both N-linked and O-linked oligosaccharides in yeast cells. In the present study, we describe the generation of a Saccharomyces cerevisiae strain capable of producing a glycoprotein with humanized Man5GlcNAc2 N-linked oligosaccharides, an intermediate of mammalian hybrid- and complex-type oligosaccharides, while suppressing O-mannosylation. First, a yeast strain that produces a glycoprotein with Man5GlcNAc2 was isolated by introducing msdS encoding α-1,2-mannosidase into a strain synthesizing Man8GlcNAc2 N-linked oligosaccharides. Next, to suppress O-mannosylation, an O-mannosyltransferase-deficient strain was generated by disrupting PMT1 and PMT2 Although the relative amount of O-linked oligosaccharides in the disruptant was reduced to approximately 40% of that in wild type cells, this strain exhibited growth defects and decreased protein productivity. To overcome the growth defects, we applied a mutagenesis technique that is based on the disparity theory of evolution. Finally, to improve protein productivity of the growth-recovered strain, vacuolar proteases PEP4 and PRB1 were further disrupted. Thus, by combining genetic engineering and disparity mutagenesis, we generated an Saccharomyces cerevisiae strain whose N- and O-linked oligosaccharide synthetic pathways were engineered to effectively produce the heterologous protein.

  17. Powder lemon juice containing oligosaccharides obtained by dextransucrase acceptor reaction synthesis and dehydrated in sprouted bed.

    PubMed

    Coelho, Raquel Macedo Dantas; Araújo, Antônia Daiana Andrade; Fontes, Cláudia Patrícia Mourão Lima; da Silva, Ana Raquel Araujo; da Costa, José Maria Correia; Rodrigues, Sueli

    2015-09-01

    Oligosaccharides can be synthesized using the sugars present in the fruit juices through the dextransucrase acceptor reaction. In the present work, the effect of reducing sugar and sucrose concentration on oligosaccharide formation in lemon juice was evaluated through response surface methodology. The oligosaccharide formation in lemon juice was favored at high concentrations of sucrose (75 g/L) and reducing sugar (75 g/L). At this synthesis conditions, an oligosaccharide concentration of 94.81 g/L was obtained with a conversion of 63.21% of the initial sugars into the target product. Oligosaccharides with degree of polymerization up to 11 were obtained. The lemon juice was dehydrated in spouted bed using maltodextrin as drying adjuvant. The powder obtained at 60°C with 20 % maltodextrin presented low moisture (2.24 %), low water activity (Aw = 0.18) and the lowest reconstitution time (~46 s). The results showed that lemon juice is suitable for oligosaccharides enzyme synthesis and can be dehydrated in spouted bed.

  18. Solution conformation of n-linked oligosaccharides

    SciTech Connect

    Homans, S.W.; Dwek, R.A.; Rademacher, T.W.

    1987-10-20

    The purpose of this review is to show how the current understanding of oligosaccharide solution conformations and dynamics has been furthered by recent developments in NMR methodology together with the use of energy calculations. First, a general strategy for the assignment of /sup 1/H NMR spectra of oligosaccharides is described. The use of these assignments for the interpretation of nuclear Overhauser effect (NOE) experiments is then discussed. The latter generate distance constraints, which, in conjunction with conformational energy calculations and molecular dynamics simulations, allow the structural and dynamic properties of a given oligosaccharide to be determined. Finally, the authors explore the extent to which the proposed recognition function of oligosaccharides can be rationalized in terms of these properties.

  19. Capillary Electrophoresis of Mono- and Oligosaccharides.

    PubMed

    Toppazzini, Mila; Coslovi, Anna; Rossi, Marco; Flamigni, Anna; Baiutti, Edi; Campa, Cristiana

    2016-01-01

    This chapter reports an overview of the recent advances in the analysis of mono- and oligosaccharides by capillary electrophoresis (CE); furthermore, relevant reviews and research articles recently published in the field are tabulated. Additionally, pretreatments and procedures applied to uncharged and acidic carbohydrates (i.e., monosaccharides and lower oligosaccharides carrying carboxylate, sulfate, or phosphate groups) are described.Representative examples of such procedures are reported in detail, upon describing robust methodologies for the study of (1) neutral oligosaccharides derivatized by reductive amination and by formation of glycosylamines; (2) sialic acid derivatized with 2-aminoacridone, released from human serum immunoglobulin G; (3) anomeric couples of neutral glycosides separated using borate-based buffers; (4) unsaturated, underivatized oligosaccharides from lyase-treated alginate.

  20. Proteomics analysis of Bacillus licheniformis in response to oligosaccharides elicitors.

    PubMed

    Reffatti, Patricia Fernanda; Roy, Ipsita; Odell, Mark; Keshavarz, Tajalli

    2014-01-01

    The role of oligosaccharides as biotic elicitors has been recognised in the enhanced production of antibiotics from fungal and bacterial cultures. The yield of bacitracin A in cultures of Bacillus licheniformis was increased after supplementation with oligoguluronate (OG), and mannan oligosaccharides (MO) and its mechanism at transcription level been established already. However, the elicitation mechanism at post transcriptional level has not been reported so far. In this paper we investigate changes in proteomics of B. licheniformis in presence of the oligosaccharide elicitors OG and MO. Differentially expressed proteins were examined using 2D-PAGE stained with colloidal Coomassie and were further identified by LC-MS/MS. We identified 19 differentially expressed proteins including those involved in carbon metabolism, energy generation, amino acid biosynthesis, oxidative and general stress response. The novel findings of this work, together with previous reports, contribute to the unravelling of the overall mechanism of elicitation in B. licheniformis cultures and reliability of the use of these elicitors for potential industrial application.

  1. Urine oligosaccharide pattern in patients with hyperprolactinaemia.

    PubMed

    Ekman, Bertil; Wahlberg, Jeanette; Landberg, Eva

    2015-11-01

    Free milk-type oligosaccharides are produced during pregnancy and lactation and may have an impact on several cells in the immune system. Our aim was to investigate if patients with isolated hyperprolactinaemia, not related to pregnancy, also have increased synthesis and urinary excretion of milk-type oligosaccharides and to compare the excretion pattern with that found during pregnancy. Urine samples were collected as morning sample from 18 patients with hyperprolactinaemia, 13 healthy controls with normal prolactin levels and four pregnant women. After purification, lactose and free oligosaccharides were analysed and quantified by high-performance anion-exchange chromatography with pulsed amperometric detection. The identity of peaks was confirmed by exoglycosidase treatment and comparison with oligosaccharide standards. Prolactin was measured in serum collected between 09 and 11 a.m. by a standardized immunochemical method. Patients with hyperprolactinaemia had higher urinary excretion of lactose than normoprolactinemic controls and urinary lactose correlated positively to prolactin levels (r = 0.51, p < 0.05). Increased levels of the fucosylated oligosaccharides 2-fucosyl lactose and lacto-di-fucotetraose were found in urine from three and two patients, respectively. The acidic oligosaccharide 3-sialyl lactose was found in high amount in urine from two patients with prolactin of >10,000 mU/l. However, pregnant women in their third trimester had the highest concentration of all these oligosaccharides and excretion increased during pregnancy. This study is first to show that both lactose and certain fucosylated and sialylated milk-type oligosaccharides are increased in some patients with hyperprolactinaemia. It remains to elucidate the functional importance of these findings.

  2. Chemoenzymatic synthesis of an isoprenoid phosphate tool for the analysis of complex bacterial oligosaccharide biosynthesis.

    PubMed

    Lujan, Donovan K; Stanziale, Jennifer A; Mostafavi, Anahita Z; Sharma, Sunita; Troutman, Jerry M

    2012-10-01

    Undecaprenyl Pyrophosphate Synthase (UPPS) is a key enzyme that catalyzes the production of bactoprenols, which act as membrane anchors for the assembly of complex bacterial oligosaccharides. One of the major hurdles in understanding the assembly of oligosaccharide assembly is a lack of chemical tools to study this process, since bactoprenols and the resulting isoprenoid-linked oligosaccharides lack handles or chromophores for use in pathway analysis. Here we describe the isolation of a new UPPS from the symbiotic microorganism Bacteroides fragilis, a key species in the human microbiome. The protein was purified to homogeneity and utilized to accept a chromophore containing farnesyl diphosphate analogue as a substrate. The analogue was utilized by the enzyme and resulted in a bactoprenyl diphosphate product with an easy to monitor tag associated with it. Furthermore, the diphosphate is shown to be readily converted to monophosphate using a common molecular biology reagent. This monophosphate product allowed for the investigation of complex oligosaccharide biosynthesis, and was used to probe the activity of glycosyltransferases involved in the well characterized Campylobacter jejuni N-linked protein glycosylation. Novel reagents similar to this will provide key tools for the study of uncharacterized oligosaccharide assemblies, and open the possibility for the development of rapid screening methodology for these biosynthetic systems.

  3. Effects of an onion by-product on bioactivity and safety markers in healthy rats.

    PubMed

    Roldán-Marín, Eduvigis; Krath, Britta N; Poulsen, Morten; Binderup, Mona-Lise; Nielsen, Tom H; Hansen, Max; Barri, Thaer; Langkilde, Søren; Cano, M Pilar; Sánchez-Moreno, Concepción; Dragsted, Lars O

    2009-12-01

    Onions are excellent sources of bioactive compounds including fructo-oligosaccharides (FOS) and polyphenols. An onion by-product was characterised in order to be developed as a potentially bioactive food ingredient. Our main aim was to investigate whether the potential health and safety effects of this onion by-product were shared by either of two derived fractions, an extract containing the onion FOS and polyphenols and a residue fraction containing mainly cell wall materials. We report here on the effects of feeding these products on markers of potential toxicity, protective enzymes and gut environment in healthy rats. Rats were fed during 4 weeks with a diet containing the products or a control feed balanced in carbohydrate. The onion by-product and the extract caused anaemia as expected in rodents for Allium products. No other toxicity was observed, including genotoxicity. Glutathione reductase (GR) and glutathione peroxidase (GPx1) activities in erythrocytes increased when rats were fed with the onion extract. Hepatic gene expression of Gr, Gpx1, catalase, 5-aminolevulinate synthase and NAD(P)H:quinone oxidoreductase was not altered in any group of the onion fed rats. By contrast, gamma-glutamate cysteine ligase catalytic subunit gene expression was upregulated but only in rats given the onion residue. The onion by-products as well as the soluble and insoluble fractions had prebiotic effects as evidenced by decreased pH, increased butyrate production and altered gut microbiota enzyme activities. In conclusion, the onion by-products have no in vivo genotoxicity, may support in vivo antioxidative defence and alter the functionality of the rat gut microbiota.

  4. Delta inulin: a novel, immunologically active, stable packing structure comprising β-d-[2 → 1] poly(fructo-furanosyl) α-d-glucose polymers

    PubMed Central

    Cooper, Peter D; Petrovsky, Nikolai

    2011-01-01

    We report a novel isoform of β-d-[2 → 1] poly(fructo-furanosyl) α-d-glucose termed delta inulin (DI), comparing it with previously described alpha (AI), beta (BI) and gamma (GI) isoforms. In vitro, DI is the most immunologically active weight/weight in human complement activation and in binding to monocytes and regulating their chemokine production and cell surface protein expression. In vivo, this translates into potent immune adjuvant activity, enhancing humoral and cellular responses against co-administered antigens. As a biocompatible polysaccharide particle, DI is safe and well tolerated by subcutaneous or intramuscular injection. Physico-chemically, DI forms as an insoluble precipitate from an aqueous solution of suitable AI, BI or GI held at 37–48°C, whereas the precipitate from the same solution at lower temperatures has the properties of AI or GI. DI can also be produced by heat conversion of GI suspensions at 56°C, whereas GI is converted from AI at 45°C. DI is distinguished from GI by its higher temperature of solution in dilute aqueous suspension and by its lower solubility in dimethyl sulfoxide, both consistent with greater hydrogen bonding in DI's polymer packing structure. DI suspensions can be dissolved by heat, re-precipitated by cooling as AI and finally re-converted back to DI by repeated heat treatment. Thus, DI, like the previously described inulin isoforms, reflects the formation of a distinct polymer aggregate packing structure via reversible noncovalent bonding. DI forms the basis for a potent new human vaccine adjuvant and further swells the growing family of carbohydrate structures with immunological activity. PMID:21147758

  5. In vitro evaluation of the fermentation properties and potential prebiotic activity of caprine cheese whey oligosaccharides in batch culture systems.

    PubMed

    Oliveira, Diana L; Costabile, Adele; Wilbey, R Andrew; Grandison, Alistair S; Duarte, Luis C; Roseiro, Luisa B

    2012-01-01

    The prebiotic effect of oligosaccharides recovered and purified from caprine whey, was evaluated by in vitro fermentation under anaerobic conditions using batch cultures at 37°C with human faeces. Effects on key gut bacterial groups were monitored over 24 h by fluorescence in situ hybridization (FISH), which was used to determine a quantitative prebiotic index score. Production of short-chain fatty acids (SCFAs) as fermentation end products was analyzed by high-performance liquid chromatography (HPLC). Growth of Bifidobacterium spp was significantly higher (P ≥ 0.05) with the purified oligosaccharides compared to the negative control. Lactic and propionic acids were the main SCFAs produced. Antimicrobial activity of the oligosaccharides was also tested, revealing no inhibition though a decrease in Staphylococcus aureus and Escherichia coli growth. These findings indicate that naturally extracted oligosaccharides from caprine whey could be used as new and valuable source of prebiotics.

  6. Filamentous fungi in good shape: microparticles for tailor-made fungal morphology and enhanced enzyme production.

    PubMed

    Driouch, Habib; Roth, Andreas; Dersch, Petra; Wittmann, Christoph

    2011-01-01

    Filamentous fungi such as Aspergillus niger are important biocatalysts for industrial production of various enzymes as well as organic acids or antibiotics. In suspended culture these microorganisms exhibit a complex morphology which typically has a strong influence on their production properties. In this regard, we have recently shown that the addition of inorganic micro particles to the culture medium is a straightforward and elegant approach to precisely tame fungal morphology. For A. niger a full range of morphological forms from pellets with different diameters to free mycelium could be adjusted by supplementation with talc powder. Aluminium oxide particles similarly affected morphology, showing that this effect is largely independent of the chemical particle composition. Exemplified for different recombinant A. niger strains enzyme production could be strongly enhanced by the addition of microparticles. This was demonstrated for the production of fructofuranosidase, an important high-value biocatalyst for pre-biotic fructo-oligosaccharides, by recombinant A. niger. In a microparticle enhanced fed-batch process, a highly productive mycelium could be achieved. The enzyme titre of 2800 U/mL finally reached was more then tenfold higher then that of any other process reported so far. Here we provide additional insights into the novel production process. This includes the confirmation of the highly selective production of the target enzyme fructofuranosidase using MALDI-TOF MS analysis. Moreover, we show that the obtained enzyme suspension can be efficiently used with minimal pre-treatment for the biosynthesis of short chain fructooligosaccharides of the inulin type, such as 1-kestose and 1-nystose, prebiotics with substantial commercial interest. In particular, these compounds are highly attractive for human consumption, since they have been shown to reduce the risk of colon cancer. In summary, the use of microparticles opens a new avenue of engineering

  7. A fingerprinting method for chondroitin/dermatan sulfate and hyaluronan oligosaccharides.

    PubMed

    Lauder, R M; Huckerby, T N; Nieduszynski, I A

    2000-04-01

    A previously published method for the analysis of glycosaminoglycan disaccharides by high pH anion exchange chromatography (Midura,R.J., Salustri,A., Calabro,A., Yanagishita,M. and Hascall,V.C. (1994), Glycobiology,4, 333-342) has been modified and calibrated for chondroitin and dermatan sulfate oligosaccharides up to hexasaccharide in size and hyaluronan oligosaccharides up to hexadecasaccharide. For hyaluronan oligosaccharides chain length controls elution position; however, for chondroitin and dermatan sulfate oligosaccharides elution times primarily depend upon the level of sulfation, although chain length and hence charge density plays a role. The sulfation position of GalNAc residues within an oligosaccharide is also important in determining its elution position. Compared to 4-sulfation a reducing terminal 6-sulfate retards elution; however, when present on an internal GalNAc residue it is the 4-sulfate containing oligosaccharide which elutes later. These effects allow discrimination between oligosaccharides differing only in the position of GalNAc sulfation. Using this simple methodology, a Dionex CarboPac PA-1 column with NaOH/NaCl eluents and detection by absorbance at 232 nm, a quantitative analytical fingerprint of a chondroitin/dermatan sulfate chain may be obtained, allowing a determination of the abundance of chondroitin sulfate, dermatan sulfate, and hyaluronan along with an analysis of structural features with a linear response to approximately 0.1 nmol. The method may readily be calibrated using either commercial disaccharides or the di- and tetrasaccharide products of a limit digest of commercial chondroitin sulfate by chondroitin ABC endolyase. Commercially available and freshly prepared shark, whale, bovine, and human cartilage chondroitin sulfates have been examined by this methodology and we have confirmed that freshly isolated shark cartilage CS contains significant amounts of the biologically important GlcA2Sbeta(1-3)GalNAc6S structure.

  8. Preparative purification of tetraantennary oligosaccharides from human asialyl orosomucoid.

    PubMed

    Stubbs, H J; Shia, M A; Rice, K G

    1997-05-01

    An approach to isolate micromole quantities of tetraantennary oligosaccharides from human orosomucoid is presented. The N-linked oligosaccharides from 500 mg of the glycoprotein were released enzymatically, desialylated, and isolated free of protein using ion exchange chromatography. The pooled oligosaccharides were converted into oligosaccharide glycosylamines by reaction with ammonium bicarbonate then coupled to BOC-tyrosine to prepare tyrosinamide oligosaccharides. These were resolved on semipreparative RP-HPLC to recover micromole quantities of six purified tyrosinamide oligosaccharides. The oligosaccharide structures were elucidated by a combination of high-field proton NMR and matrix-assisted time of flight mass spectrometry and included biantennary, triantennary, monofucosylated triantennary, tetraantennary, monofucosylated tetraantennary, and a tetraantennary containing a single polylactosamine extension. Edman degradation was utilized to reverse the tyrosinamide oligosaccharide derivatization leading to the generation of reducing oligosaccharides. These were used to characterize the elution profile of asialyl orosomucoid oligosaccharides on high pH anion exchange chromatography. This application of tyrosinamide derivatization has allowed for the first time the complete resolution of the complex oligosaccharide mixture from orosomucoid on a semipreparative scale in a single chromatogram and provide the first NMR characterization of polylactosamine tetraantennary oligosaccharide from this substrate. This study demonstrates the broad utility of the tyrosinamide derivatization to develop oligosaccharide libraries useful for probing the biological functions of glycosylation.

  9. Comprehensive peptidomic and glycomic evaluation reveals that sweet whey permeate from colostrum is a source of milk protein-derived peptides and oligosaccharides

    PubMed Central

    Dallas, David C.; Weinborn, Valerie; de Moura Bell, Juliana M.L.N.; Wang, Meng; Parker, Evan A.; Guerrero, Andres; Hettinga, Kasper A.; Lebrilla, Carlito B.; German, J. Bruce; Barile, Daniela

    2014-01-01

    Whey permeate is a co-product obtained when cheese whey is passed through an ultrafiltration membrane to concentrate whey proteins. Whey proteins are retained by the membrane, whereas the low-molecular weight compounds such as lactose, salts, oligosaccharides and peptides pass through the membrane yielding whey permeate. Research shows that bovine milk from healthy cows contains hundreds of naturally occurring peptides – many of which are homologous with known antimicrobial and immunomodulatory peptides – and nearly 50 oligosaccharide compositions (not including structural isomers). As these endogenous peptides and oligosaccharides have low-molecular weight and whey permeate is currently an under-utilized product stream of the dairy industry, we hypothesized that whey permeate may serve as an inexpensive source of naturally occurring functional peptides and oligosaccharides. Laboratory fractionation of endogenous peptides and oligosaccharides from bovine colostrum sweet whey was expanded to pilot-scale. The membrane fractionation methodology used was similar to the methods commonly used industrially to produce whey protein concentrate and whey permeate. Pilot-scale fractionation was compared to laboratory-scale fractionation with regard to the identified peptides and oligosaccharide compositions. Results were interpreted on the basis of whether industrial whey permeate could eventually serve as a source of functional peptides and oligosaccharides. The majority (96%) of peptide sequences and the majority (96%) of oligosaccharide compositions found in the laboratory-scale process were mirrored in the pilot-scale process. Moreover, the pilot-scale process recovered an additional 33 peptides and 1 oligosaccharide not identified from the laboratory-scale extraction. Both laboratory- and pilot-scale processes yielded peptides deriving primarily from the protein β-casein. The similarity of the laboratory-and pilot-scale's resulting peptide and oligosaccharide

  10. Comprehensive peptidomic and glycomic evaluation reveals that sweet whey permeate from colostrum is a source of milk protein-derived peptides and oligosaccharides.

    PubMed

    Dallas, David C; Weinborn, Valerie; de Moura Bell, Juliana M L N; Wang, Meng; Parker, Evan A; Guerrero, Andres; Hettinga, Kasper A; Lebrilla, Carlito B; German, J Bruce; Barile, Daniela

    2014-09-01

    Whey permeate is a co-product obtained when cheese whey is passed through an ultrafiltration membrane to concentrate whey proteins. Whey proteins are retained by the membrane, whereas the low-molecular weight compounds such as lactose, salts, oligosaccharides and peptides pass through the membrane yielding whey permeate. Research shows that bovine milk from healthy cows contains hundreds of naturally occurring peptides - many of which are homologous with known antimicrobial and immunomodulatory peptides - and nearly 50 oligosaccharide compositions (not including structural isomers). As these endogenous peptides and oligosaccharides have low-molecular weight and whey permeate is currently an under-utilized product stream of the dairy industry, we hypothesized that whey permeate may serve as an inexpensive source of naturally occurring functional peptides and oligosaccharides. Laboratory fractionation of endogenous peptides and oligosaccharides from bovine colostrum sweet whey was expanded to pilot-scale. The membrane fractionation methodology used was similar to the methods commonly used industrially to produce whey protein concentrate and whey permeate. Pilot-scale fractionation was compared to laboratory-scale fractionation with regard to the identified peptides and oligosaccharide compositions. Results were interpreted on the basis of whether industrial whey permeate could eventually serve as a source of functional peptides and oligosaccharides. The majority (96%) of peptide sequences and the majority (96%) of oligosaccharide compositions found in the laboratory-scale process were mirrored in the pilot-scale process. Moreover, the pilot-scale process recovered an additional 33 peptides and 1 oligosaccharide not identified from the laboratory-scale extraction. Both laboratory- and pilot-scale processes yielded peptides deriving primarily from the protein β-casein. The similarity of the laboratory-and pilot-scale's resulting peptide and oligosaccharide profiles

  11. Xyloglucan oligosaccharides promote growth and activate cellulase: Evidence for a role of cellulase in cell expansion. [Pisum sativum L

    SciTech Connect

    McDougall, G.J.; Fry, S.C. )

    1990-07-01

    Oligosaccharides produced by the action of fungal cellulase on xyloglucans promoted the elongation of etiolated pea (Pisum sativum L.) stem segments in a straight-growth bioassay designed for the determination of auxins. The oligosaccharides were most active at about 1 micromolar. We tested the relative growth-promoting activities of four HPLC-purified oligosaccharides which shared a common glucose{sub 4} {center dot} xylose{sub 3} (XG7) core. The substituted oligosaccharides XG8 (glucose{sub 4} {center dot} xylose{sub 3} {center dot} galactose) and XG9n (glucose{sub 4} {center dot} xylose{sub 3} {center dot} galactose{sub 2}) were more effective than XG7 itself and XG9 (glucose{sub 4} {center dot} xylose{sub 3} {center dot} galactose {center dot} fucose). The same oligosaccharides also promoted the degradation, assayed viscometrically, of xyloglucan by an acidic cellulase from bean (Phaseolus vulgaris L.) leaves. The oligosaccharides were highly active at 10{sup {minus}4} molar, causing up to a fourfold increase in activity, but the effect was still detectable at 1 micromolar. Those oligosaccharides (XG8 and XG9n) which best promoted growth, stimulated cellulase activity to the greatest extent. The oligosaccharides did not stimulate the action of the cellulase in an assay based on the conversion of ({sup 3}H)xyloglucan to ethanol-soluble fragments. This suggests that the oligosaccharides enhanced the midchain hydrolysis of xyloglucan molecules (which would rapidly reduce the viscosity of the solution), at the expense of cleavage near the termini (which would yield ethanol-soluble products).

  12. Oligosaccharide Binding in Escherichia coli Glycogen Synthase

    SciTech Connect

    Sheng, Fang; Yep, Alejandra; Feng, Lei; Preiss, Jack; Geiger, James H.

    2010-11-17

    Glycogen/starch synthase elongates glucan chains and is the key enzyme in the synthesis of glycogen in bacteria and starch in plants. Cocrystallization of Escherichia coli wild-type glycogen synthase (GS) with substrate ADPGlc and the glucan acceptor mimic HEPPSO produced a closed form of GS and suggests that domain-domain closure accompanies glycogen synthesis. Cocrystallization of the inactive GS mutant E377A with substrate ADPGlc and oligosaccharide results in the first oligosaccharide-bound glycogen synthase structure. Four bound oligosaccharides are observed, one in the interdomain cleft (G6a) and three on the N-terminal domain surface (G6b, G6c, and G6d). Extending from the center of the enzyme to the interdomain cleft opening, G6a mostly interacts with the highly conserved N-terminal domain residues lining the cleft of GS. The surface-bound oligosaccharides G6c and G6d have less interaction with enzyme and exhibit a more curled, helixlike structural arrangement. The observation that oligosaccharides bind only to the N-terminal domain of GS suggests that glycogen in vivo probably binds to only one side of the enzyme to ensure unencumbered interdomain movement, which is required for efficient, continuous glucan-chain synthesis.

  13. Human milk oligosaccharides: only the breast.

    PubMed

    McVeagh, P; Miller, J B

    1997-08-01

    Over 100 years ago it was first deduced that a major component of human milk must be an unidentified carbohydrate that was not found in cows milk. At first this was thought to be a form of lactose and was called gynolactose. We now know that this was not a single carbohydrate but a complex mixture of approximately 130 different oligosaccharides. Although small amounts of a few oligosaccharides have been found in the milk of other mammals, this rich diversity of sugars is unique to human milk. The oligosaccharide content of human milk varies with the infant's gestation, the duration of lactation, diurnally and with the genetic makeup of the mother. Milk oligosaccharides have a number of functions that may protect the health of the breast fed infant. As they are not digested in the small intestine, they form the 'soluble' fibre of breast milk and their intact structure is available to act as competitive ligands protecting the breast-fed infant from pathogens. There is a growing list of pathogens for which a specific oligosaccharide ligand has been described in human milk. They are likely to form the model for future therapeutic and prophylactic anti-microbials. They provide substrates for bacteria in the infant colon and thereby contribute to the difference in faecal pH and faecal flora between breast and formula-fed infants. They may also be important as a source of sialic acid, essential for brain development.

  14. Partly Fermented Infant Formulae With Specific Oligosaccharides Support Adequate Infant Growth and Are Well-Tolerated

    PubMed Central

    Huet, Frédéric; Abrahamse-Berkeveld, Marieke; Tims, Sebastian; Simeoni, Umberto; Beley, Gérard; Savagner, Christoph; Vandenplas, Yvan; Hourihane, Jonathan O’B.

    2016-01-01

    ABSTRACT Objective: Fermented formulae (FERM) and a specific mixture of 90% short-chain galacto-oligosaccharides and 10% long-chain fructo-oligosaccharides (scGOS/lcFOS; 9:1) have a potential beneficial effect on gastrointestinal function and microbiota development in infants. The present study assessed the safety and tolerance of the combination of partly fermented infant milk formulae and scGOS/lcFOS compared with either 1 feature, in healthy term infants. Methods: Four hundred thirty-two infants were enrolled before 28 days of age and followed up to 17 weeks of age and assigned to 1 of the 4 groups: (i) formula with scGOS/lcFOS, (ii) scGOS/lcFOS + 15% FERM, (iii) scGOS/lcFOS + 50% FERM, or (iv) 50% fermented formula (50% FERM). Primary outcome was daily weight gain during intervention (equivalence criterion: difference in daily weight gain ≤3 g/day). Infants’ anthropometrics, formula intake, number, and type of (serious) AEs were monitored monthly. Stool samples were collected at baseline and after 17 weeks for analysis of physiological and microbiological parameters. Results: Equivalence of weight gain per day was demonstrated in both the intention-to-treat and per-protocol population, with a mean weight gain (SD) of 29.7 (6.1), 28.2 (4.8), 28.5 (5.0), and 28.7 (5.9) g/day for the groups i to iv respectively. No differences were observed in other growth parameters, formula intake, and the number or severity of AEs. In all scGOS/lcFOS-containing formulae, a beneficial effect of scGOS/lcFOS was observed, indicated by the lower pH, lower Clostridium difficile levels, and higher secretory immunoglobulin A levels. Conclusions: The partly fermented infant milk formulae containing the specific mixture scGOS/lcFOS were well-tolerated and resulted in normal growth in healthy infants. PMID:27472478

  15. Structures and application of oligosaccharides in human milk

    PubMed Central

    KOBATA, Akira

    2010-01-01

    Comparative study of the oligosaccharide profiles of individual human milk revealed the presence of three different patterns. Four oligosaccharides containing the Fucα1-2Gal group were missing in the milk of non-secretor, and three oligosaccharides containing the Fucα1-4GlcNAc group were missing in the milk of Lewis negative individuals. Disappearance of some major oligosaccharides in these samples led to the finding of five novel minor oligosaccharides, which were hidden under the missing oligosaccharides. Following these studies, structures of many novel milk oligosaccharides were elucidated. At least 13 core oligosaccharides were found in these oligosaccharides. By adding α-fucosyl residues and sialic acid residues to these core oligosaccharides, more than one hundred oligosaccharides were formed. All these oligosaccharides contain lactose at their reducing termini. This evidence, together with the deletion phenomena found in the milk oligosaccharides of non-secretor and Lewis negative individuals, suggested that the oligosaccharides are formed from lactose by the concerted action of glycosyltransferases, which are responsible for elongation and branching of the Galβ1-4GlcNAc group in the sugar chains of glycoconjugates on the surface of epithelial cells. Therefore, oligosaccharides in human milk could include many structures, starting from the Galβ1-4GlcNAc group in the sugar chains of various glycoconjugates. Many lines of evidence recently indicated that virulent enteric bacteria and viruses start their infection by binding to particular sugar chains of glycoconjugates on the target cell surfaces. Therefore, milk oligosaccharides could be useful for developing drugs, which inhibit the infection of bacteria and viruses. PMID:20689231

  16. Mode of action of xylogalacturonan hydrolase towards xylogalacturonan and xylogalacturonan oligosaccharides

    PubMed Central

    2004-01-01

    XGH (xylogalacturonan hydrolase; GH 28) is an enzyme that is capable of degrading XGA (xylogalacturonan), which is a polymer of α-D-galacturonic acid, highly substituted with β-D-xylose. XGA is present in cell walls of various plants and exudates, such as gum tragacanth. XGA oligosaccharides were derived from an XGH digestion of gum tragacanth, then fractionated, and analysed for their sugar composition and structure by matrix-assisted laser-desorption ionization–time-of-flight MS and nanospray MS. Several oligosaccharides from XGA were identified with different galacturonic acid/xylose ratios including five oligosaccharide isomers. Although XGH can act as an endo-enzyme, product-progression profiling showed that the disaccharide GalAXyl was predominantly produced from XGA by XGH, which indicated also an exolytic action. The latter was further supported by degradation studies of purified oligosaccharide GalA4Xyl3. It was shown that XGH acted from the non-reducing end towards the reducing end of this oligosaccharide, and showed the processive character of XGH. The results from this study further show that although XGH prefers to act between two xylosidated GalA units, it tolerates unsubstituted GalA units in its −1 and +1 subsites. PMID:15560751

  17. Uptake of chitosan-derived D-glucosamine oligosaccharides in Streptomyces coelicolor A3(2).

    PubMed

    Viens, Pascal; Dubeau, Marie-Pierre; Kimura, Akane; Desaki, Yoshitake; Shinya, Tomonori; Shibuya, Naoto; Saito, Akihiro; Brzezinski, Ryszard

    2015-05-01

    The csnR gene, localized at the beginning of an operon, csnR-K, which organization is conserved through many actinomycete genomes, was previously shown to repress the transcription of the chitosanase gene csnA in Streptomyces lividans. However, knowledge on the function of the whole csnR-K operon in the metabolism of chitosan (an N-deacetylated derivative of chitin) remained limited. Mutants of S. coelicolor A3(2) harboring partial or total deletions of the csnR-K operon were analyzed for their capacity to uptake glucosamine oligosaccharides (GlcN)n. The csnR-K operon was autoregulated by CsnR repressor and its transcription was inducible by GlcN oligosaccharides. The operon controlled the uptake of GlcN oligosaccharides in S. coelicolor A3(2), with a minor contribution to the consumption of monomeric GlcN but not chitin-related N-acetylated derivatives. The deletion of the whole operon abolished the uptake of GlcN oligosaccharides. The CsnEFG transporter encoded by this operon is the front door for the assimilation of chitosan-derived hydrolysis products in S. coelicolor A3(2). The ATP-binding component MsiK was essential for CsnEFG transport function. Also, deletion of msiK abolished the induction of csnA transcription by GlcN oligosaccharides.

  18. Influence of the physico-chemical characteristics of chito-oligosaccharides (COS) on antioxidant activity.

    PubMed

    Mengíbar, Marian; Mateos-Aparicio, Inmaculada; Miralles, Beatriz; Heras, Angeles

    2013-09-12

    Chito-oligosaccharides (COS) are being used as important functional materials for many applications due to their bioactivities. The aim of this research has been to assess the relationship between the physico-chemical characteristics, average molecular weight (Mw), acetylation degree (DA), polymerization degree (DP) and specially sequence composition determined by MALDI-TOF MS and the antioxidant properties of COS. These oligosaccharides were obtained by enzymatic depolymerization with chitosanase and lysozyme using a specific chitosan and its reacetylated product. The COS fraction below 5 kDa obtained from chitosanase depolymerization showed the highest capacity to scavenge DPPH radicals and to reduce Fe(3+). A correlation was found between the relative amount of molecules with a given A/D (acetylated vs deacetylated units) ratio within the COS and their antioxidant activity, which could be used to predict the antioxidant behavior of a fraction of chito-oligosaccharides.

  19. On-line separation and characterization of hyaluronan oligosaccharides derived from radical depolymerization

    PubMed Central

    Zhao, Xue; Yang, Bo; Li, Lingyun; Zhang, Fuming; Linhardt, Robert J.

    2013-01-01

    Hydroxyl radicals are widely implicated in the oxidation of carbohydrates in biological and industrial processes and are often responsible for their structural modification resulting in functional damage. In this study, the radical depolymerization of the polysaccharide hyaluronan was studied in a reaction with hydroxyl radicals generated by Fenton Chemistry. A simple method for isolation and identification of the resulting non-sulfated oligosaccharide products of oxidative depolymerization was established. Hyaluronan oligosaccharides were analyzed using ion-pairing reversed phase high performance liquid chromotography coupled with tandem electrospray mass spectrometry. The sequence of saturated hyaluronan oligosaccharides having even- and odd-numbers of saccharide units, afforded through oxidative depolymerization, were identified. This study represents a simple, effective ‘fingerprinting’ protocol for detecting the damage done to hyaluronan by oxidative radicals. This study should help reveal the potential biological outcome of reactive-oxygen radical-mediated depolymerization of hyaluronan. PMID:23768593

  20. Detection of N-(1-deoxy-d-fructos-1-yl) Fumonisins B2 and B3 in Corn by High-Resolution LC-Orbitrap MS

    PubMed Central

    Matsuo, Yosuke; Takahara, Kentaro; Sago, Yuki; Kushiro, Masayo; Nagashima, Hitoshi; Nakagawa, Hiroyuki

    2015-01-01

    The existence of glucose conjugates of fumonisin B2 (FB2) and fumonisin B3 (FB3) in corn powder was confirmed for the first time. These “bound-fumonisins” (FB2 and FB3 bound to glucose) were identified as N-(1-deoxy-d-fructos-1-yl) fumonisin B2 (NDfrc-FB2) and N-(1-deoxy-d-fructos-1-yl) fumonisin B3 (NDfrc-FB3) respectively, based on the accurate mass measurements of characteristic ions and fragmentation patterns using high-resolution liquid chromatography-Orbitrap mass spectrometry (LC-Orbitrap MS) analysis. Treatment on NDfrc-FB2 and NDfrc-FB3 with the o-phthalaldehyde (OPA) reagent also supported that d-glucose binding to FB2 and FB3 molecules occurred to their primary amine residues. PMID:26389955

  1. Mixed-Linkage Glucan Oligosaccharides Produced by Automated Glycan Assembly Serve as Tools To Determine the Substrate Specificity of Lichenase.

    PubMed

    Dallabernardina, Pietro; Schuhmacher, Frank; Seeberger, Peter H; Pfrengle, Fabian

    2017-03-02

    The mixed-linkage (1→3),(1→4)-d-glucan (MLG) specific glycosyl hydrolase lichenase is an important biochemical tool for the structural characterization of MLGs. It holds potential for application in the brewery, animal feed, and biofuel industries. Several defined MLG oligosaccharides obtained by automated glycan assembly are used to analyze the substrate specificities of Bacillus subtilis lichenase. Two glucose building blocks (BBs), equipped with a temporary fluorenylmethyloxycarbonyl chloride (Fmoc) protecting group in the C-3 or C-4 position, served to assemble different oligosaccharides by using an automated oligosaccharide synthesizer. Light-induced cleavage of the glycan products from the solid support followed by global deprotection provided seven MLG oligosaccharides of different length and connectivity. After incubation of the MLG oligosaccharides with lichenase, the digestion products were analyzed by HPLC-MS. These digestion experiments provided insights into the enzyme's active site that is in line with other recent evidence suggesting that the substrate specificity of lichenases has to be reconsidered. These results demonstrate that synthetic MLG oligosaccharides are useful tools to analyze mixed-linkage β-glucanases.

  2. New Food Oligosaccharides via Alternansucrase Acceptor Reactions

    USDA-ARS?s Scientific Manuscript database

    Alternansucrase [EC 2.4.1.140] is a glycosyltransferase from food-grade bacteria that is capable of synthesizing unique polysaccharides and oligosaccharides from sucrose. The enzyme typically produces the high-molecular weight polysaccharide alternan. However, in the presence of low-molecular weig...

  3. The Cytosolic Oligosaccharide-Degrading Proteome of Butyrivibrio Proteoclasticus

    PubMed Central

    Dunne, Jonathan C.; Kelly, William J.; Leahy, Sinead C.; Li, Dong; Bond, Judy J.; Peng, Lifeng; Attwood, Graeme T.; Jordan, T. William

    2015-01-01

    The growth and productivity of ruminants depends on a complex microbial community found in their fore-stomach (rumen), which is able to breakdown plant polysaccharides and ferment the released sugars. Butyrivibrio proteoclasticus B316T is a Gram-positive polysaccharide-degrading, butyrate-producing bacterium that is present at high numbers in the rumen of animals consuming pasture or grass silage based diets. B316T is one of a small number of rumen fibrolytic microbes capable of efficiently degrading and utilizing xylan, as well as being capable of utilizing arabinose, xylose, pectin and starch. We have therefore carried out a proteomic analysis of B316T to identify intracellular enzymes that are implicated in the metabolism of internalized xylan. Three hundred and ninety four proteins were identified including enzymes that have potential to metabolize assimilated products of extracellular xylan digestion. Identified enzymes included arabinosidases, esterases, an endoxylanase, and β-xylosidase. The presence of intracellular debranching enzymes indicated that some hemicellulosic side-chains may not be removed until oligosaccharides liberated by extracellular digestion have been assimilated by the cells. The results support a model of extracellular digestion of hemicellulose to oligosaccharides that are then transported to the cytoplasm for further digestion by intracellular enzymes. PMID:28248275

  4. Active Site Mapping of Xylan-Deconstructing Enzymes with Arabinoxylan Oligosaccharides Produced by Automated Glycan Assembly.

    PubMed

    Senf, Deborah; Ruprecht, Colin; de Kruijff, Goswinus H M; Simonetti, Sebastian O; Schuhmacher, Frank; Seeberger, Peter H; Pfrengle, Fabian

    2017-03-02

    Xylan-degrading enzymes are crucial for the deconstruction of hemicellulosic biomass, making the hydrolysis products available for various industrial applications such as the production of biofuel. To determine the substrate specificities of these enzymes, we prepared a collection of complex xylan oligosaccharides by automated glycan assembly. Seven differentially protected building blocks provided the basis for the modular assembly of 2-substituted, 3-substituted, and 2-/3-substituted arabino- and glucuronoxylan oligosaccharides. Elongation of the xylan backbone relied on iterative additions of C4-fluorenylmethoxylcarbonyl (Fmoc) protected xylose building blocks to a linker-functionalized resin. Arabinofuranose and glucuronic acid residues have been selectively attached to the backbone using fully orthogonal 2-(methyl)naphthyl (Nap) and 2-(azidomethyl)benzoyl (Azmb) protecting groups at the C2 and C3 hydroxyls of the xylose building blocks. The arabinoxylan oligosaccharides are excellent tools to map the active site of glycosyl hydrolases involved in xylan deconstruction. The substrate specificities of several xylanases and arabinofuranosidases were determined by analyzing the digestion products after incubation of the oligosaccharides with glycosyl hydrolases. © 2017 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

  5. Novel process for the coproduction of xylo-oligosaccharides, fermentable sugars, and lignosulfonates from hardwood.

    PubMed

    Huang, Caoxing; Jeuck, Ben; Du, Jing; Yong, Qiang; Chang, Hou-Min; Jameel, Hasan; Phillips, Richard

    2016-11-01

    Many biorefineries have not been commercialized due to poor economic returns from final products. In this work, a novel process has been developed to coproduce valuable sugars, xylo-oligosaccharides, and lignosulfonates from hardwood. The modified process includes a mild autohydrolysis pretreatment, which enables for the recovery of the xylo-oligosaccharides in auto-hydrolysate. Following enzymatic hydrolysis, the residue is sulfomethylated to produce lignosulfonates. Recycling the sulfomethylation residues increased both the glucan recovery and lignosulfonate production. The glucose recovery was increased from 81.7% to 87.9%. Steady state simulation using 100g of hardwood produced 46.7g sugars, 5.9g xylo-oligosaccharides, and 25.7g lignosulfonates, which were significantly higher than that produced from the no-recycling process with 39.1g sugars, 5.9g xylo-oligosaccharides, and 15.0g lignosulfonates. The results indicate that this novel biorefinery process can improve the production of fermentable sugars and lignosulfonate from hardwood as compared to a conventional biorefinery process.

  6. Inhibition of the aggregation of lactoferrin and (-)-epigallocatechin gallate in the presence of polyphenols, oligosaccharides, and collagen peptide.

    PubMed

    Yang, Wei; Liu, Fuguo; Xu, Chenqi; Sun, Cuixia; Yuan, Fang; Gao, Yanxiang

    2015-05-27

    The aggregation of lactoferrin and (-)-epigallocatechin gallate (EGCG) was inhibited by polyphenols, oligosaccharides, and collagen peptide in this study. Polyphenols, oligosaccharides, or collagen peptide can effectively prevent the formation of lactoferrin-EGCG aggregates, respectively. The addition sequence of lactoferrin, polyphenols (oligosaccharides or collagen peptide) and EGCG can affect the turbidity and particle size of the ternary complexes in the buffer solution; however, it hardly affected the ζ-potential and fluorescence characteristics. With either positive or negative charge, polyphenols and collagen peptide disrupted the formation of lactoferrin-EGCG aggregate mainly through the mechanism of its competition with EGCG molecules which surrounded the lactoferrin molecule surface with weaker binding affinities, forming polyphenols or a collagen peptide-lactoferrin-EGCG ternary complex; for neutral oligosaccharides, the ternary complex was generated mainly through steric effects, accompanied by a change in the lactoferrin secondary structure induced by gallic acid, chlorogenic acid, and xylo-oligosaccharide. Polyphenols, oligosaccharides, or collagen peptide restraining the formation of lactoferrin-EGCG aggregate could be applied in the design of clear products in the food, pharmaceutical, and cosmetic industries.

  7. Evolutionary Glycomics: Characterization of Milk Oligosaccharides in Primates

    PubMed Central

    Tao, Nannan; Wu, Shuai; Kim, Jaehan; An, Hyun Joo; Hinde, Katie; Power, Michael L.; Gagneux, Pascal; German, J. Bruce; Lebrilla, Carlito B.

    2011-01-01

    Free oligosaccharides are abundant components of mammalian milk and have primary roles as prebiotic compounds, in immune defense, and in brain development. Mass spectrometry-based technique is applied to profile milk oligosaccharides from apes (chimpanzee, gorilla, and siamang), new world monkeys (golden lion tamarin and common marmoset), and an old world monkey (rhesus). The purpose of this study was to evaluate the patterns of primate milk oligosaccharide composition from a phylogenetic perspective in order to assess the extent to which the compositions of hMOs derives from ancestral, primate patterns as opposed to more recent evolutionary events. Milk oligosaccharides were quantitated by nanoflow liquid chromatography on chip-based devices. The relative abundances of fucosylated and sialylated milk oligosaccharides in primates were also determined. For a systematic and comprehensive study of evolutionary patterns of milk oligosaccharides, cluster analysis of primate milk was performed using the chromatographic profile. In general, the oligosaccharides in primate milk, including humans, are more complex and exhibit greater diversity compared to the ones in non-primate milk. A detailed comparison of the oligosaccharides across evolution revealed non-sequential developmental pattern, i.e. that primate milk oligosaccharides do not necessarily cluster according to the primate phylogeny. This report represents the first comprehensive and quantitative effort to profile and elucidate the structures of free milk oligosaccharides so that they can be related to glycan function in different primates. PMID:21214271

  8. Blackbody Infrared Radiative Dissociation of Protonated Oligosaccharides

    NASA Astrophysics Data System (ADS)

    Fentabil, Messele A.; Daneshfar, Rambod; Kitova, Elena N.; Klassen, John S.

    2011-12-01

    The dissociation pathways, kinetics, and energetics of protonated oligosaccharides in the gas phase were investigated using blackbody infrared radiative dissociation (BIRD). Time-resolved BIRD measurements were performed on singly protonated ions of cellohexaose (Cel6), which is composed of β-(1 → 4)-linked glucopyranose rings, and five malto-oligosaccharides (Malx, where x = 4-8), which are composed of α-(1 → 4)-linked glucopyranose units. At the temperatures investigated (85-160 °C), the oligosaccharides dissociate at the glycosidic linkages or by the loss of a water molecule to produce B- or Y-type ions. The Y ions dissociate to smaller Y or B ions, while the B ions yield exclusively smaller B ions. The sequential loss of water molecules from the smallest B ions (B1 and B2) also occurs. Rate constants for dissociation of the protonated oligosaccharides and the corresponding Arrhenius activation parameters (Ea and A) were determined. The Ea and A-factors measured for protonated Malx (x > 4) are indistinguishable within error (~19 kcal mol-1, 1010 s-1), which is consistent with the ions being in the rapid energy exchange limit. In contrast, the Arrhenius parameters for protonated Cel6 (24 kcal mol-1, 1012 s-1) are significantly larger. These results indicate that both the energy and entropy changes associated with the glycosidic bond cleavage are sensitive to the anomeric configuration. Based on the results of this study, it is proposed that formation of B and Y ions occurs through a common dissociation mechanism, with the position of the proton establishing whether a B or Y ion is formed upon glycosidic bond cleavage.

  9. Human milk oligosaccharides: the novel modulator of intestinal microbiota.

    PubMed

    Jeong, Kyunghun; Nguyen, Vi; Kim, Jaehan

    2012-08-01

    Human milk, which nourishes the early infants, is a source of bioactive components for the infant growth, development and commensal formulation as well. Human milk oligosaccharide is a group of complex and diverse glycans that is apparently not absorbed in human gastrointestinal tract. Although most mammalian milk contains oligosaccharides, oligosaccharides in human milk exhibit unique features in terms of their types, amounts, sizes, and functionalities. In addition to the prevention of infectious bacteria and the development of early immune system, human milk oligosaccharides are able to facilitate the healthy intestinal microbiota. Bifidobacterial intestinal microbiota appears to be established by the unilateral interaction between milk oligosaccharides, human intestinal activity and commensals. Digestibility, membrane transportation and catabolic activity by bacteria and intestinal epithelial cells, all of which are linked to the structural of human milk oligosaccharides, are crucial in determining intestinal microbiota.

  10. Divergent Synthesis of Heparan Sulfate Oligosaccharides

    PubMed Central

    2015-01-01

    Heparan sulfates are implicated in a wide range of biological processes. A major challenge in deciphering their structure and activity relationship is the synthetic difficulties to access diverse heparan sulfate oligosaccharides with well-defined sulfation patterns. In order to expedite the synthesis, a divergent synthetic strategy was developed. By integrating chemical synthesis and two types of O-sulfo transferases, seven different hexasaccharides were obtained from a single hexasaccharide precursor. This approach combined the flexibility of chemical synthesis with the selectivity of enzyme-catalyzed sulfations, thus simplifying the overall synthetic operations. In an attempt to establish structure activity relationships of heparan sulfate binding with its receptor, the synthesized oligosaccharides were incorporated onto a glycan microarray, and their bindings with a growth factor FGF-2 were examined. The unique combination of chemical and enzymatic approaches expanded the capability of oligosaccharide synthesis. In addition, the well-defined heparan sulfate structures helped shine light on the fine substrate specificities of biosynthetic enzymes and confirm the potential sequence of enzymatic reactions in biosynthesis. PMID:26574650

  11. Healing efficiency of oligosaccharides generated from almond gum (Prunus amygdalus) on dermal wounds of adult rats.

    PubMed

    Bouaziz, Fatma; Ben Romdhane, Molka; Boisset Helbert, Claire; Buon, Laurine; Bhiri, Fatma; Bardaa, Sana; Driss, Dorra; Koubaa, Mohamed; Fakhfakh, Akram; Sahnoun, Zouhair; Kallel, Fatma; Zghal, Najiba; Ellouz Chaabouni, Semia

    2014-08-01

    Almond gum is a naturally occurring polymer produced by almond trees and shrubs. Its abundance, as well as its low cost production makes it a potential feedstock for use in food and pharmaceuticals. In this regard, almond gum oligosaccharides were enzymatically generated, purified and their monosaccharide composition assessed using gas chromatography-flame ionization detector. Oligosaccharide analyses show that the most prominent residues were galactose and arabinose with traces of xylose, rhamnose, glucose and mannose. The glycosyl linkage positions were analyzed using gas chromatography - mass spectrometry showing a main chain composed of galactose units [→3)-Gal-(1→] branched mainly with arabinose residues [Ara-(1→]. The potent role of the generated oligosaccharides on rats wound healing was investigated. They have been applied either alone or supplemented, as active substance, with cream formulation, on full-thickness wound created on the dorsum of the rats. The effect of oligosaccharides was assessed by measuring the wound closure percentage, reaching an average of around 100% when applied alone or supplemented to cream formulation. The healing percentage for the control group was only 74.3% at the same day. The histological evaluation of skin sections visualized by light microscopy revealed an improved collagen deposition and an increased fibroblast and vascular densities. Copyright © 2014 Tissue Viability Society. Published by Elsevier Ltd. All rights reserved.

  12. Preparation, characterisation and use for antioxidant oligosaccharides of a cellulase from abalone (Haliotis discus hannai) viscera.

    PubMed

    Tao, Zhi-Peng; Sun, Le-Chang; Qiu, Xu-Jian; Cai, Qiu-Feng; Liu, Guang-Ming; Su, Wen-Jin; Cao, Min-Jie

    2016-07-01

    In China, abalone (Haliotis discus hannai) production is growing annually. During industrial processing, the viscera, which are abundant of cellulase, are usually discarded or processed into low-value feedstuff. Thus, it is of interest to obtain cellulase from abalone viscera and investigate its application for preparation of functional oligosaccharides. A cellulase was purified from the hepatopancreas of abalone by ammonium sulfate precipitation and two-steps column chromatography. The molecular weight of the cellulase was 45 kDa on SDS-PAGE. Peptide mass fingerprinting analysis yielded 103 amino acid residues, which were identical to cellulases from other species of abalone. Substrate specificity analysis indicated that the cellulase is an endo-1,4-β-glucanase. Hydrolysis of seaweed Porphyra haitanensis polysaccharides by the enzyme produced oligosaccharides with degree of polymerisation of two to four, whose monosaccharide composition was 58% galactose, 4% glucose and 38% xylose. The oligosaccharides revealed 2,2'-diphenyl-1-picrylhydrazyl free radical as well as hydrogen peroxide scavenging activity. It is feasible and meaningful to utilise cellulase from the viscera of abalone for preparation of functional oligosaccharides. © 2015 Society of Chemical Industry. © 2015 Society of Chemical Industry.

  13. Sodium alginate oligosaccharides attenuate hypertension and associated kidney damage in Dahl salt-sensitive rats fed a high-salt diet.

    PubMed

    Terakado, Shouko; Ueno, Mai; Tamura, Yuki; Toda, Natsuko; Yoshinaga, Mariko; Otsuka, Kie; Numabe, Atsushi; Kawabata, Yukari; Murota, Itsuki; Sato, Nobuyuki; Uehara, Yoshio

    2012-01-01

    In this article, the antihypertensive effects of sodium alginate oligosaccharides, enzymatic products of high molecular natural alginate from sea weeds, in Dahl salt-sensitive (Dahl S) rats were investigated. Dahl S rats fed a high-salt (4% NaCl) diet were treated with sodium alginate oligosaccharides (4% or 8% w/w) for 7 weeks. Systolic blood pressure (SBP) was measured by the tail-cuff method, and hypertensive cardiovascular benefits and kidney damage were assessed. Glomerular function and morphological sclerosis were determined. SBP increased in an age-dependent manner in the untreated Dahl S rats. Sodium alginate oligosaccharide treatment attenuated the increase in SBP in a dose-dependent manner. The heart and aortic walls weighed less in the rats treated with sodium alginate oligosaccharides than in the untreated rats. The SBP reduction was associated with a decrease in urinary protein excretion and an increase in the creatinine clearance rate. Sodium alginate oligosaccharides significantly attenuated hypertensive glomerular sclerosis and arterial injury in the kidney. Fractional excretion of sodium (FENa) decreased in low-salt Dahl S rats and increased with a salt challenge. The alginate oligosaccharides decreased FENa in high-salt Dahl S rats. The results of this study suggest that sodium alginate oligosaccharides attenuate salt-induced hypertension in Dahl S rats. This reduction is associated with decreases in cardiovascular and renal damage.

  14. Improved liquid chromatography-MS/MS of heparan sulfate oligosaccharides via chip-based pulsed makeup flow.

    PubMed

    Huang, Yu; Shi, Xiaofeng; Yu, Xiang; Leymarie, Nancy; Staples, Gregory O; Yin, Hongfeng; Killeen, Kevin; Zaia, Joseph

    2011-11-01

    Microfluidic chip-based hydrophilic interaction chromatography (HILIC) is a useful separation system for liquid chromatography-mass spectrometry (LC-MS) in compositional profiling of heparan sulfate (HS) oligosaccharides; however, ions observed using HILIC LC-MS are low in charge. Tandem MS of HS oligosaccharide ions with low charge results in undesirable losses of SO(3) from precursor ions during collision induced dissociation. One solution is to add metal cations to stabilize sulfate groups. Another is to add a nonvolatile, polar compound such as sulfolane, a molecule known to supercharge proteins, to produce a similar effect for oligosaccharides. We demonstrate use of a novel pulsed makeup flow (MUF) HPLC-chip. The chip enables controlled application of additives during specified chromatographic time windows and thus minimizes the extent to which nonvolatile additives build up in the ion source. The pulsed MUF system was applied to LC-MS/MS of HS oligosaccharides. Metal cations and sulfolane were tested as additives. The most promising results were obtained for sulfolane, for which supercharging of the oligosaccharide ions increased their signal strengths relative to controls. Tandem MS of these supercharged precursor ions showed decreased abundances of product ions from sulfate losses yet more abundant product ions from backbone cleavages.

  15. The principal fucosylated oligosaccharides of human milk exhibit prebiotic properties on cultured infant microbiota

    PubMed Central

    Yu, Zhuo-Teng; Chen, Ceng; Kling, David E; Liu, Bo; McCoy, John M; Merighi, Massimo; Heidtman, Matthew; Newburg, David S

    2013-01-01

    Breast-fed infant microbiota is typically rich in bifidobacteria. Herein, major human milk oligosaccharides (HMOS) are assessed for their ability to promote the growth of bifidobacteria and to acidify their environment, key features of prebiotics. During in vitro anaerobic fermentation of infant microbiota, supplementation by HMOS significantly decreased the pH even greater than supplementation by fructooligosaccharide (FOS), a prebiotic positive control. HMOS elevated lactate concentrations, increased the proportion of Bifidobacterium spp. in culture, and through their fermentation into organic acids, decreased the proportion of Escherichia and Clostridium perfringens. Three principal components of HMOS, 2′-fucosyllactose, lactodifucotetraose and 3-fucosyllactose, were consumed in these cultures. These three principal oligosaccharides of human milk were then individually tested as supplements for in vitro growth of four individual representative strains of infant gut microbes. Bifidobacterium longum JCM7007 and B. longum ATCC15697 efficiently consumed oligosaccharides and produced abundant lactate and short-chain fatty acids, resulting in significant pH reduction. The specificity of fermentation differed by microbe species and strain and by oligosaccharide structure. Escherichia coli K12 and C. perfringens did not utilize appreciable fucosylated oligosaccharides, and a typical mixture of organic acid fermentation products inhibited their growth. In summary, 2′-fucosyllactose, lactodifucotetraose, and 3-fucosyllactose, when cultured with B. longum JCM7007 and B. longum ATCC15697, exhibit key characteristics of a prebiotic in vitro. If these bifidobacteria are representative of pioneering or keystone species for human microbiota, fucosylated HMOS could strongly promote colonization and maintenance of a mutualist symbiotic microbiome. Thus, these simple glycans could mediate beneficial effects of human milk on infant health. PMID:23028202

  16. The principal fucosylated oligosaccharides of human milk exhibit prebiotic properties on cultured infant microbiota.

    PubMed

    Yu, Zhuo-Teng; Chen, Ceng; Kling, David E; Liu, Bo; McCoy, John M; Merighi, Massimo; Heidtman, Matthew; Newburg, David S

    2013-02-01

    Breast-fed infant microbiota is typically rich in bifidobacteria. Herein, major human milk oligosaccharides (HMOS) are assessed for their ability to promote the growth of bifidobacteria and to acidify their environment, key features of prebiotics. During in vitro anaerobic fermentation of infant microbiota, supplementation by HMOS significantly decreased the pH even greater than supplementation by fructooligosaccharide (FOS), a prebiotic positive control. HMOS elevated lactate concentrations, increased the proportion of Bifidobacterium spp. in culture, and through their fermentation into organic acids, decreased the proportion of Escherichia and Clostridium perfringens. Three principal components of HMOS, 2'-fucosyllactose, lactodifucotetraose and 3-fucosyllactose, were consumed in these cultures. These three principal oligosaccharides of human milk were then individually tested as supplements for in vitro growth of four individual representative strains of infant gut microbes. Bifidobacterium longum JCM7007 and B. longum ATCC15697 efficiently consumed oligosaccharides and produced abundant lactate and short-chain fatty acids, resulting in significant pH reduction. The specificity of fermentation differed by microbe species and strain and by oligosaccharide structure. Escherichia coli K12 and C. perfringens did not utilize appreciable fucosylated oligosaccharides, and a typical mixture of organic acid fermentation products inhibited their growth. In summary, 2'-fucosyllactose, lactodifucotetraose, and 3-fucosyllactose, when cultured with B. longum JCM7007 and B. longum ATCC15697, exhibit key characteristics of a prebiotic in vitro. If these bifidobacteria are representative of pioneering or keystone species for human microbiota, fucosylated HMOS could strongly promote colonization and maintenance of a mutualist symbiotic microbiome. Thus, these simple glycans could mediate beneficial effects of human milk on infant health.

  17. Influence of glycosidic linkages and molecular weight on the fermentation of maltose-based oligosaccharides by human gut bacteria.

    PubMed

    Sanz, María Luz; Côté, Gregory L; Gibson, Glenn R; Rastall, Robert A

    2006-12-27

    A structure-function study was carried out to increase knowledge of how glycosidic linkages and molecular weights of carbohydrates contribute toward the selectivity of fermentation by gut bacteria. Oligosaccharides with maltose as the common carbohydrate source were used. Potentially prebiotic alternansucrase and dextransucrase maltose acceptor products were synthesized and separated into different molecular weights using a Bio-gel P2 column. These fractions were characterized by matrix-assisted laser desorption/ionization time-of-flight. Nonprebiotic maltooligosaccharides with degrees of polymerization (DP) from three to seven were commercially obtained for comparison. Growth selectivity of fecal bacteria on these oligosaccharides was studied using an anaerobic in vitro fermentation method. In general, carbohydrates of DP3 showed the highest selectivity towards bifidobacteria; however, oligosaccharides with a higher molecular weight (DP6-DP7) also resulted in a selective fermentation. Oligosaccharides with DPs above seven did not promote the growth of "beneficial" bacteria. The knowledge of how specific structures modify the gut microflora could help to find new prebiotic oligosaccharides.

  18. Subcutaneous administration of sodium alginate oligosaccharides prevents salt-induced hypertension in Dahl salt-sensitive rats.

    PubMed

    Moriya, Chikako; Shida, Yui; Yamane, Yuki; Miyamoto, Yuki; Kimura, Midori; Huse, Naomi; Ebisawa, Kaori; Kameda, Yuki; Nishi, Ayaka; Du, DongDong; Yoshinaga, Mariko; Murota, Itsuki; Sato, Nobuyuki; Uehara, Yoshio

    2013-01-01

    We investigated the mechanism of antihypertensive effects of sodium alginate oligosaccharides, which are enzymatic products of high-molecular-weight natural alginate from seaweeds, in Dahl salt-sensitive (Dahl S) rats. Dahl S rats fed a high-salt (4% NaCl) diet were subcutaneously administered sodium alginate oligosaccharides (60 mg/day using a continuous osmotic mini-pump) for 14 days. Systolic blood pressure (SBP) was measured using the tail-cuff method, and we determined the influence of the alginate treatment on the metabolism of sodium by measuring sodium excretions in the feces and urine. SBP increased in an age-dependent manner in the untreated Dahl S rats. Sodium alginate oligosaccharide treatment via the subcutaneous route almost completely abolished salt-induced hypertension in Dahl S rats fed a high-salt diet. The level of fecal or urinary sodium excretion did not significantly change during the treatment period with the alginate oligosaccharides. The reduction in SBP rapidly recovered after cessation of the treatment. Moreover, the level of urinary protein excretion was lower in the treated Dahl S rats than in the untreated rats during the experimental period. Our results suggest that sodium alginate oligosaccharides attenuate salt-induced hypertension in Dahl S rats not through reducing salt absorption, but probably through a direct action on vascular vessels.

  19. Of the milk sugars, galactose, but not prebiotic galacto-oligosaccharide, improves insulin sensitivity in male Sprague-Dawley rats.

    PubMed

    Stahel, Priska; Kim, Julie J; Xiao, Changting; Cant, John P

    2017-01-01

    Consumption of dairy products reduces risk of type 2 diabetes. Milk proteins and fats exhibit anti-diabetic properties but milk sugars have been studied little in this context. Galactose from milk lactose is readily converted to glycogen in the liver but its effects on insulin sensitivity have not been assessed. Prebiotic oligosaccharides from milk alter gut microbiota and can thereby influence host metabolism. Our objective was to assess the effect on insulin sensitivity of dietary galactose compared to glucose and fructose, and fermentable galacto-oligosaccharides compared to non-fermentable methylcellulose. Diets containing 15% of dry matter from glucose, fructose, galactose, galacto-oligosaccharides, or methylcellulose were fed to 36 rats per diet for 9 weeks. Hyperinsulinemic-euglycemic clamps with [3-3H]glucose infusion and a steady-state 2-[1-14C]deoxyglucose bolus injection were used to assess insulin sensitivity and glucose uptake indices. Tissue was collected in fed, fasted and fasted, insulin-stimulated states. Galactose increased glucose infusion rate during the clamp by 53% and decreased endogenous glucose production by 57% compared to glucose and fructose. Fed-state hepatic glycogen content was greater with galactose compared to glucose and fructose, consistent with a potentiation of the insulin effect on glycogen synthase by dephosphorylation. Galactose decreased the fecal Firmicutes:Bacteroidetes ratio while galacto-oligosaccharides increased abundance of fecal Bifidobacterium spp. 481-fold compared to methylcellulose, and also increased abundance of Lactobacillus spp. and Bacteroidetes. Galacto-oligosaccharides did not affect glucose infusion rate or endogenous glucose production during basal or clamp periods compared to methylcellulose. Galactose at 15% of daily intake improved hepatic insulin sensitivity in rats compared to glucose and fructose. Galactose caused an increase in fed-state hepatic glycogen content and a favourable shift in gut

  20. Of the milk sugars, galactose, but not prebiotic galacto-oligosaccharide, improves insulin sensitivity in male Sprague-Dawley rats

    PubMed Central

    Kim, Julie J.; Xiao, Changting; Cant, John P.

    2017-01-01

    Background Consumption of dairy products reduces risk of type 2 diabetes. Milk proteins and fats exhibit anti-diabetic properties but milk sugars have been studied little in this context. Galactose from milk lactose is readily converted to glycogen in the liver but its effects on insulin sensitivity have not been assessed. Prebiotic oligosaccharides from milk alter gut microbiota and can thereby influence host metabolism. Our objective was to assess the effect on insulin sensitivity of dietary galactose compared to glucose and fructose, and fermentable galacto-oligosaccharides compared to non-fermentable methylcellulose. Methods Diets containing 15% of dry matter from glucose, fructose, galactose, galacto-oligosaccharides, or methylcellulose were fed to 36 rats per diet for 9 weeks. Hyperinsulinemic-euglycemic clamps with [3-3H]glucose infusion and a steady-state 2-[1-14C]deoxyglucose bolus injection were used to assess insulin sensitivity and glucose uptake indices. Tissue was collected in fed, fasted and fasted, insulin-stimulated states. Results Galactose increased glucose infusion rate during the clamp by 53% and decreased endogenous glucose production by 57% compared to glucose and fructose. Fed-state hepatic glycogen content was greater with galactose compared to glucose and fructose, consistent with a potentiation of the insulin effect on glycogen synthase by dephosphorylation. Galactose decreased the fecal Firmicutes:Bacteroidetes ratio while galacto-oligosaccharides increased abundance of fecal Bifidobacterium spp. 481-fold compared to methylcellulose, and also increased abundance of Lactobacillus spp. and Bacteroidetes. Galacto-oligosaccharides did not affect glucose infusion rate or endogenous glucose production during basal or clamp periods compared to methylcellulose. Conclusions Galactose at 15% of daily intake improved hepatic insulin sensitivity in rats compared to glucose and fructose. Galactose caused an increase in fed-state hepatic glycogen

  1. Nitric oxide-releasing chitosan oligosaccharides as antibacterial agents.

    PubMed

    Lu, Yuan; Slomberg, Danielle L; Schoenfisch, Mark H

    2014-02-01

    Secondary amine-functionalized chitosan oligosaccharides of different molecular weights (i.e., ~2500, 5000, 10,000) were synthesized by grafting 2-methyl aziridine from the primary amines on chitosan oligosaccharides, followed by reaction with nitric oxide (NO) gas under basic conditions to yield N-diazeniumdiolate NO donors. The total NO storage, maximum NO flux, and half-life of the resulting NO-releasing chitosan oligosaccharides were controlled by the molar ratio of 2-methyl aziridine to primary amines (e.g., 1:1, 2:1) and the functional group surrounding the N-diazeniumdiolates (e.g., polyethylene glycol (PEG) chains), respectively. The secondary amine-modified chitosan oligosaccharides greatly increased the NO payload over existing biodegradable macromolecular NO donors. In addition, the water-solubility of the chitosan oligosaccharides enabled their penetration across the extracellular polysaccharides matrix of Pseudomonas aeruginosa biofilms and association with embedded bacteria. The effectiveness of these chitosan oligosaccharides at biofilm eradication was shown to depend on both the molecular weight and ionic characteristics. Low molecular weight and cationic chitosan oligosaccharides exhibited rapid association with bacteria throughout the entire biofilm, leading to enhanced biofilm killing. At concentrations resulting in 5-log killing of bacteria in Pseudomonas aeruginosa (P. aeruginosa) biofilms, the NO-releasing and control chitosan oligosaccharides elicited no significant cytotoxicity to mouse fibroblast L929 cells in vitro. Copyright © 2013 Elsevier Ltd. All rights reserved.

  2. Nitric Oxide-Releasing Chitosan Oligosaccharides as Antibacterial Agents

    PubMed Central

    Lu, Yuan; Slomberg, Danielle L.; Schoenfisch, Mark H.

    2014-01-01

    Secondary amine-functionalized chitosan oligosaccharides of different molecular weights (i.e., ~2500, 5000, 10000) were synthesized by grafting 2-methyl aziridine from the primary amines on chitosan oligosaccharides, followed by reaction with nitric oxide (NO) gas under basic conditions to yield N-diazeniumdiolate NO donors. The total NO storage, maximum NO flux, and half-life of the resulting NO-releasing chitosan oligosaccharides were controlled by the molar ratio of 2-methyl aziridine to primary amines (e.g., 1:1, 2:1) and the functional group surrounding the N-diazeniumdiolates (e.g., polyethylene glycol (PEG) chains), respectively. The secondary amine-modified chitosan oligosaccharides greatly increased the NO payload over existing biodegradable macromolecular NO donors. In addition, the water-solubility of the chitosan oligosaccharides enabled their penetration across the extracellular polysaccharides matrix of Pseudomonas aeruginosa biofilms and association with embedded bacteria. The effectiveness of these chitosan oligosaccharides at biofilm eradication was shown to depend on both the molecular weight and ionic characteristics. Low molecular weight and cationic chitosan oligosaccharides exhibited rapid association with bacteria throughout the entire biofilm, leading to enhanced biofilm killing. At concentrations resulting in 5-log killing of bacteria in Pseudomonas aeruginosa biofilms, the NO-releasing and control chitosan oligosaccharides elicited no significant cytotoxicity to mouse fibroblast L929 cells in vitro. PMID:24268196

  3. Consumption of human milk oligosaccharides by gut-related microbes.

    PubMed

    Marcobal, Angela; Barboza, Mariana; Froehlich, John W; Block, David E; German, J Bruce; Lebrilla, Carlito B; Mills, David A

    2010-05-12

    Human milk contains large amounts of complex oligosaccharides that putatively modulate the intestinal microbiota of breast-fed infants by acting as decoy binding sites for pathogens and as prebiotics for enrichment of beneficial bacteria. Several bifidobacterial species have been shown to grow well on human milk oligosaccharides. However, few data exist on other bacterial species. This work examined 16 bacterial strains belonging to 10 different genera for growth on human milk oligosaccharides. For this propose, a chemically defined medium, ZMB1, was used, which allows vigorous growth of a number of gut-related microorganisms in a fashion similar to complex media. Interestingly, Bifidobacterium longum subsp. infantis, Bacteroides fragilis , and Bacteroides vulgatus strains were able to metabolize milk oligosaccharides with high efficiency, whereas Enterococcus , Streptococcus , Veillonella , Eubacterium , Clostridium , and Escherichia coli strains grew less well or not at all. Mass spectrometry-based glycoprofiling of the oligosaccharide consumption behavior revealed a specific preference for fucosylated oligosaccharides by Bi. longum subsp. infantis and Ba. vulgatus. This work expands the current knowledge of human milk oligosaccharide consumption by gut microbes, revealing bacteroides as avid consumers of this substrate. These results provide insight on how human milk oligosaccharides shape the infant intestinal microbiota.

  4. Oligosaccharides in human milk: structural, functional, and metabolic aspects.

    PubMed

    Kunz, C; Rudloff, S; Baier, W; Klein, N; Strobel, S

    2000-01-01

    Research on human milk oligosaccharides (HMOs) has received much attention in recent years. However, it started about a century ago with the observation that oligosaccharides might be growth factors for a so-called bifidus flora in breast-fed infants and extends to the recent finding of cell adhesion molecules in human milk. The latter are involved in inflammatory events recognizing carbohydrate sequences that also can be found in human milk. The similarities between epithelial cell surface carbohydrates and oligosaccharides in human milk strengthen the idea that specific interactions of those oligosaccharides with pathogenic microorganisms do occur preventing the attachment of microbes to epithelial cells. HMOs may act as soluble receptors for different pathogens, thus increasing the resistance of breast-fed infants. However, we need to know more about the metabolism of oligosaccharides in the gastrointestinal tract. How far are oligosaccharides degraded by intestinal enzymes and does oligosaccharide processing (e.g. degradation, synthesis, and elongation of core structures) occur in intestinal epithelial cells? Further research on HMOs is certainly needed to increase our knowledge of infant nutrition as it is affected by complex oligosaccharides.

  5. Consumption of Human Milk Oligosaccharides by Gut-related Microbes

    PubMed Central

    Marcobal, Angela; Barboza, Mariana; Froehlich, John W.; Block, David E.; German, J. Bruce; Lebrilla, Carlito B.; Mills, David A.

    2010-01-01

    Human milk contains large amounts of complex oligosaccharides that putatively modulate the intestinal microbiota of breast-fed infants by acting as decoy binding sites for pathogens and as prebiotics for enrichment of beneficial bacteria. Several bifidobacterial species have been shown to grow well on human milk oligosaccharides. However, little data exists on other bacterial species. In this work we examined 16 bacterial strains belonging to 10 different genera for growth on human milk oligosaccharides. For this propose, we used a chemically-defined medium, ZMB1, which allows vigorous growth of a number gut–related microorganisms in a fashion similar to complex media. Interestingly, Bifidobacterium longum subsp. infantis, Bacteroides fragilis and Bacteroides vulgatus strains were able to metabolize milk oligosaccharides with high efficiency, while Enterococcus, Streptococcus, Veillonella, Eubacterium, Clostridium, and Escherichia coli strains grew less well or not at all. Mass spectrometry-based glycoprofiling of the oligosaccharide consumption behavior revealed a specific preference for fucosylated oligosaccharides by Bifidobacterium longum subsp. infantis and Bacteroides vulgatus. This work expands the current knowledge of human milk oligosaccharides consumption by gut microbes, revealing bacteroides as avid consumer of this substrate. These results provide insight on how human milk oligosaccharides shape the infant intestinal microbiota. PMID:20394371

  6. In vitro fermentation characteristics of selected oligosaccharides by swine fecal microflora.

    PubMed

    Smiricky-Tjardes, M R; Flickinger, E A; Grieshop, C M; Bauer, L L; Murphy, M R; Fahey, G C

    2003-10-01

    The objective of this study was to quantify the fermentation characteristics of oligosaccharides present in feed ingredients or isolated for dietary supplementation. Substrates studied included short-chain fructooligosaccharides, medium-chain fructooligosaccharides, long-chain fructooligosaccharides, raffinose, stachyose, soy solubles, granular and liquid forms of transgalactooligosaccharides, glucooligosaccharides, mannanoligosaccharides, and xylooligosaccharides. Three healthy pigs that had never received antibiotics served as sources of fecal inoculum. Each substrate was fermented in vitro; samples were taken at 0, 2, 4, 8, and 12 h, and pH change and short-chain fatty acid (SCFA) and gas production determined. Gas production at 12 h did not differ (P > 0.05) among all fructooligosaccharides, transgalactooligosaccharides, soy solubles, and xylooligosaccharides. Raffinose, stachyose, and raffinose + stachyose fermentation resulted in the greatest (P < 0.05) gas production at 12 h of all substrates tested. The rate of gas production was greatest (P < 0.05) for stachyose and least (P < 0.05) for glucooligosaccharides and mannanoligosaccharides. Substrate did not affect (P > 0.05) time to attain maximal rate of gas production. The pH at 12 h for all fructooligosaccharides and xylooligosaccharides did not differ (P > 0.05). The pH values at 12 h for raffinose, stachyose, and raffinose + stachyose were highest (P < 0.05) compared with all other substrates. Total SCFA production at 12 h was similar for all fructooligosaccharides and transgalactooligosaccharides, glucooligosaccharides, and soy solubles. Total SCFA production was greatest (P < 0.05) for xylooligosaccharides, stachyose, and raffinose + stachyose, and least (P < 0.05) for mannanoligosaccharides and raffinose. Stachyose fermentation resulted in the greatest (P < 0.05) rate and earliest time to attain maximal rate of SCFA production. All oligosaccharides studied were readily fermentable but varied in amount

  7. Metabolism and biological functions of human milk oligosaccharides.

    PubMed

    Bertino, E; Peila, C; Giuliani, F; Martano, C; Cresi, F; Di Nicola, P; Occhi, L; Sabatino, G; Fabris, C

    2012-01-01

    It is well known that breastfeeding is beneficial both for its nutritional properties and for the presence of biologically active compounds. Among these, human milk oligosaccharides (HMOs), representing the third largest fraction of human milk, have been assigned important biological functions, such as prebiotic and immunomodulatory and antimicrobial effects. HMOs are synthesized in the mammary gland by glycosyltransferase enzymes and can be divided in core-oligosaccharides, sialo-oligosaccharides, fucosyl-oligosaccharides and sialo-fucosyl-oligosaccharides on the basis of their chemical structure. Glycosyltransferases enzymes are partially regulated by genetic mechanisms; according to the expression of secretory and Lewis' genes, it is possible to classify human milk in 4 different secretory groups. We hereby present a review of the current knowledge concerning HMOs, their metabolism and main biological functions.

  8. Inhibition of N-linked oligosaccharide trimming does not interfere with surface expression of certain integral membrane proteins.

    PubMed Central

    Burke, B; Matlin, K; Bause, E; Legler, G; Peyrieras, N; Ploegh, H

    1984-01-01

    The effects of 1-deoxynojirimycin (dNM) and 1-deoxymannojirimycin (dMM), inhibitors of oligosaccharide trimming glucosidase I and mannosidase I, respectively, on the biosynthesis of vesicular stomatitis virus G protein, influenza virus hemagglutinin, and human class I histocompatibility antigens were investigated. Although the oligosaccharides of these membrane glycoproteins were greatly altered, neither dNM nor dMM interferred with their surface expression, as determined by a variety of assays, including accessibility to proteases and antibodies; neither did these drugs inhibit production of infectious virus particles. Images Fig. 1. Fig. 2. Fig. 3. Fig. 4. PMID:6232135

  9. Separation of anionic oligosaccharides by high-performance liquid chromatography

    SciTech Connect

    Green, E.D.; Baenziger, J.U.

    1986-10-01

    The authors have developed methods for rapid fractionation of anionic oligosaccharides containing sulfate and/or sialic acid moieties by high-performance liquid chromatography (HPLC). Ion-exchange HPLC on amine-bearing columns (Micropak AX-10 and AX-5) at pH 4.0 is utilized to separate anionic oligosaccharides bearing zero, one, two, three, or four charges, independent of the identity of the anionic moieties (sulfate and/or sialic acid). Ion-exchange HPLC at pH 1.7 allows separation of neutral, mono-, di-, and tetrasialylated, monosulfated, and disulfated oligosaccharides. Oligosaccharides containing three sialic acid residues and those bearing one each of sulfate and sialic acid, however, coelute at pH 1.7. Since the latter two oligosaccharide species separate at pH 4.0, analysis at pH 4.0 followed by analysis at pH 1.7 can be utilized to completely fractionate complex mixtures of sulfated and sialylated oligosaccharides. Ion-suppression amine adsorption HPLC has previously been shown to separate anionic oligosaccharides on the basis of net carbohydrate content (size). In this study they demonstrate the utility of ion-suppression amine adsorption HPLC for resolving sialylated oligosaccharide isomers which differ only in the linkages of sialic acid residues (..cap alpha..2,3 vs ..cap alpha..2,6) and/or location of ..cap alpha..2,3- and ..cap alpha..2,6-linked sialic acid moieties on the peripheral branches of oligosaccharides. These two methods can be used in tandem to separate oligosaccharides, both analytically and preparatively, based on their number, types, and linkages of anionic moieties.

  10. Structures of sulfated oligosaccharides in human trachea mucin glycoproteins.

    PubMed

    Sangadala, S; Bhat, U R; Mendicino, J

    1993-09-08

    The structures of high molecular weight sulfated oligosaccharide chains in mucins purified from the sputum of a patient with cystic fibrosis and blood group H determinant were established. Reduced oligosaccharides released by treatment with alkaline borohydride were separated by ion exchange chromatography on DEAE-Agarose and a fraction containing multisulfated chains was further purified by lectin affinity chromatography to completely remove small amounts of sialylated chains. A major sulfated oligosaccharide fraction containing chains with an average of 160 to 200 sugar residues was isolated by gel filtration on BioGel P-10 columns and individual subfractions were characterized by methylation analysis, periodate oxidation and sequential glycosidase digestion before and after desulfation. Carbohydrate analysis yielded Fuc, Gal and GldNAc in a ratio of 1:2:2.1 and only one galactosaminitol residue for every 160- to 200 sugar residues. The average molecular weight of oligosaccharide chains in these fractions was between 27,000 and 40,000 daltons. Structural analysis showed that these high molecular weight chains contained varying amounts of the repeating unit shown in the following oligosaccharide. Only one in about every 10 repeating units contained sulfate esters. Several shorter chains which contain 2 to 3 sulfate esters were also isolated from this multisulfated oligosaccharide fraction. The structures proposed for these oligosaccharides indicate that they are lower molecular weight chains with the same general structure as those found in the high molecular weight sulfated oligosaccharides. Taken collectively, the results of these studies show that a major sulfated oligosaccharide fraction in respiratory mucin purified from the mucus of patients with cystic fibrosis contains high molecular weight branched chains that consist of a repeating oligosaccharide sequence with sulfate linked to the 6 positions of galactose and possibly GlcNAc residues in the side chains.

  11. O-oligosaccharidyl-1-amino-1-deoxyalditols as intermediates for fluorescent labelling of oligosaccharides.

    PubMed

    Miller, Janice G; Farkas, Vladimír; Sharples, Sandra C; Fry, Stephen C

    2007-01-15

    Reducing monosaccharides were efficiently converted to stable 1-amino-1-deoxyalditols (=glycamines; distinguished from glycosylamines by mass-spectrometry) during incubation at 20 degrees C in saturated aqueous NH(4)HCO(3) containing NaCNBH(3). Potentially useful by-products included a novel, fully-reduced dimer (the corresponding secondary glycamine) and several relatively long-lived, unreduced products. With increasing incubation time, monomers exceeded dimers. Reducing disaccharides and oligosaccharides underwent similar reactions at their reducing termini; the yield of dimers decreased with increasing oligosaccharide M(r). The O-oligosaccharidyl-1-amino-1-deoxyalditols (OADs) obtained by reductive amination of oligosaccharides reacted readily with lissamine rhodamine sulfonyl chloride to yield OAD-sulforhodamine conjugates linked by a stable sulfonamide bond. Conditions for this reaction were optimised (borate buffer, pH9.0-9.5). The highly fluorescent OAD-sulforhodamine products were purified on a C(18) cartridge. They were electrophoretically immobile at pH2.0 and 6.5, and migrated towards the anode in borate buffer, pH9.4. The OAD-sulforhodamines were amenable to TLC and were excellent substrates for enzymic transglycosylation and for glycosylhydrolase action.

  12. Synthesis and purification of galacto-oligosaccharides: state of the art.

    PubMed

    Vera, Carlos; Córdova, Andrés; Aburto, Carla; Guerrero, Cecilia; Suárez, Sebastián; Illanes, Andrés

    2016-12-01

    Lactose-derived non-digestible oligosaccharides are prominent components of functional foods. Among them, galacto-oligosaccharides (GOS) outstand for being prebiotics whose health-promoting effects are supported on strong scientific evidences, having unique properties as substitutes of human milk oligosaccharides in formulas for newborns and infants. GOS are currently produced enzymatically in a kinetically-controlled reaction of lactose transgalactosylation catalyzed by β-galactosidases from different microbial strains. The enzymatic synthesis of GOS, although being an established technology, still offers many technological challenges and opportunities for further development that has to be considered within the framework of functional foods which is the most rapidly expanding market within the food sector. This paper presents the current technological status of GOS production, its main achievements and challenges. Most of the problems yet to be solved refer to the rather low GOS yields attainable that rarely exceed 40 %, corresponding to lactose conversions around 60 %. This means that the product or reaction (raw GOS) contains significant amounts of residual lactose and monosaccharides (glucose and galactose). Efforts to increase such yields have been for the most part unsuccessful, even though improvements by genetic and protein engineering strategies are to be expected in the near future. Low yields impose a burden on downstream processing to obtain a GOS product of the required purity. Different strategies for raw GOS purification are reviewed and their technological significance is appraised.

  13. Multifunctional fructans and raffinose family oligosaccharides

    PubMed Central

    den Ende, Wim Van

    2013-01-01

    Fructans and raffinose family oligosaccharides (RFOs) are the two most important classes of water-soluble carbohydrates in plants. Recent progress is summarized on their metabolism (and regulation) and on their functions in plants and in food (prebiotics, antioxidants). Interest has shifted from the classic inulin-type fructans to more complex fructans. Similarly, alternative RFOs were discovered next to the classic RFOs. Considerable progress has been made in the understanding of structure–function relationships among different kinds of plant fructan metabolizing enzymes. This helps to understand their evolution from (invertase) ancestors, and the evolution and role of so-called “defective invertases.” Both fructans and RFOs can act as reserve carbohydrates, membrane stabilizers and stress tolerance mediators. Fructan metabolism can also play a role in osmoregulation (e.g., flower opening) and source–sink relationships. Here, two novel emerging roles are highlighted. First, fructans and RFOs may contribute to overall cellular reactive oxygen species (ROS) homeostasis by specific ROS scavenging processes in the vicinity of organellar membranes (e.g., vacuole, chloroplasts). Second, it is hypothesized that small fructans and RFOs act as phloem-mobile signaling compounds under stress. It is speculated that such underlying antioxidant and oligosaccharide signaling mechanisms contribute to disease prevention in plants as well as in animals and in humans. PMID:23882273

  14. Analysis of Milk Oligosaccharides by Mass Spectrometry.

    PubMed

    Wu, Lauren D; Ruhaak, L Renee; Lebrilla, Carlito B

    2017-01-01

    Human milk oligosaccharides (HMOs) are a highly abundant constituent in human milk, and its protective and prebiotic properties have attracted considerable attention. HMOs have been shown to directly and indirectly benefit the overall health of the infant due to a number of functions including serving as a beneficial food for gut bacteria, block to pathogens, and aiding in brain development. Researchers are currently exploring whether these structures may act as possible disease and nutrition biomarkers. Because of this, rapid-throughput methods are desired to investigate biological activity in large patient sets. We have optimized a rapid-throughput protocol to analyze human milk oligosaccharides using micro-volumes of human breast milk for nutritional biomarkers. This method may additionally be applied to other biological fluid substrates such as plasma, urine, and feces. The protocol involves lipid separation via centrifugation, protein precipitation using ethanol, alditol reduction with sodium borohydride, and a final solid-phase extraction purification step using graphitized carbon cartridges. Samples are analyzed using HPLC-Chip/TOF-MS and data filtered on Agilent MassHunter using an in-house library. Individual structural identification is matched against a previously developed HMO library using accurate mass and retention time. Using this method will allow in-depth characterization and profiling of HMOs in large patient sets, and will ease the process of discovering significant nutritional biomarkers in human milk.

  15. Mannan Oligosaccharides in Nursery Pig Nutrition and Their Potential Mode of Action

    PubMed Central

    Halas, Veronika; Nochta, Imre

    2012-01-01

    Simple Summary The aim of the paper is to provide a review of mannan oligosaccharide products in relation to their growth promoting effect and mode of action. Mannan oligosaccharide products maintain intestinal integrity and the digestive and absorptive function of the gut in the post-weaning period in pigs and enhance disease resistance by promoting antigen presentation. We find that dietary supplementation has growth promoting effects in pigs kept in a poor hygienic environment, while the positive effect of MOS is not observed in healthy pig herds with high hygienic standards. Abstract Mannan oligosaccharides (MOSs) are often referred to as one of the potential alternatives for antimicrobial growth promoters. The aim of the paper is to provide a review of mannan oligosaccharide products in relation to their growth promoting effect and mode of action based on the latest publications. We discuss the dietary impact of MOSs on (1) microbial changes, (2) morphological changes of gut tissue and digestibility of nutrients, and (3) immune response of pigs after weaning. Dietary MOSs maintain the intestinal integrity and the digestive and absorptive function of the gut in the post-weaning period. Recent results suggest that MOS enhances the disease resistance in swine by promoting antigen presentation facilitating thereby the shift from an innate to an adaptive immune response. Accordingly, dietary MOS supplementation has a potential growth promoting effect in pigs kept in a poor hygienic environment, while the positive effect of MOS is not observed in healthy pig herds with high hygienic standards that are able to maintain a high growth rate after weaning. PMID:26486920

  16. Effect of dual-type oligosaccharides on constipation in loperamide-treated rats

    PubMed Central

    Han, Sung Hee; Hong, Ki Bae; Kim, Eun Young; Ahn, So Hyun

    2016-01-01

    BACKGROUND/OBJECTIVES Constipation is a condition that can result from intestinal deformation. Because humans have an upright posture, the effects of gravity can cause this shape deformation. Oligosaccharides are common prebiotics and their effects on bowel health are well known. However, studies of the physiological functionality of a product that contains both lactulose and galactooligosaccharides are insufficient. We investigated the constipation reduction effect of a dual-type oligosaccharide, Dual-Oligo, in loperamide-treated rats. MATERIALS/METHODS Dual-Oligo consists of galactooligosaccharides (15.80%) and lactulose (51.67%). Animals were randomly divided into four groups, the normal group (normal), control group (control), low concentration of Dual-Oligo (LDO) group, and high concentration of Dual-Oligo (HDO) group. After 7 days of oral administration, fecal pellet amount, fecal weight, water content of fecal were measured. Blood chemistry, short-chain fatty acid (SCFA), gastrointestinal transit ratio and length and intestinal mucosa were analyzed. RESULTS Dual-Oligo increased the fecal weight, and water content of feces in rats with loperamide-induced constipation. Gastrointestinal transit ratio and length and area of intestinal mucosa significantly increased after treatment with Dual-Oligo in loperamide-induced rats. A high concentration of Dual-Oligo tended to produce more acetic acid than that observed for the control group, and Dual-Oligo affected the production of total SCFA. Bifidobacteria concentration of cecal contents in the high-concentration oligosaccharide (HDO) and low-concentration oligosaccharide (LDO) groups was similar to the result of the normal group. CONCLUSIONS These results showed that Dual-Oligo is a functional material that is derived from a natural food product and is effective in ameliorating constipation. PMID:27909555

  17. Extending Synthetic Routes for Oligosaccharides by Enzyme, Substrate and Reaction Engineering

    NASA Astrophysics Data System (ADS)

    Seibel, Jürgen; Jördening, Hans-Joachim; Buchholz, Klaus

    The integration of all relevant tools for bioreaction engineering has been a recent challenge. This approach should notably favor the production of oligo- and polysaccharides, which is highly complex due to the requirements of regio- and stereoselectivity. Oligosaccharides (OS) and polysaccharides (PS) have found many interests in the fields of food, pharmaceuticals, and cosmetics due to different specific properties. Food, sweeteners, and food ingredients represent important sectors where OS are used in major amounts. Increasing attention has been devoted to the sophisticated roles of OS and glycosylated compounds, at cell or membrane surfaces, and their function, e.g., in infection and cancer proliferation. The challenge for synthesis is obvious, and convenient approaches using cheap and readily available substrates and enzymes will be discussed. We report on new routes for the synthesis of oligosaccharides (OS), with emphasis on enzymatic reactions, since they offer unique properties, proceeding highly regio- and stereoselective in water solution, and providing for high yields in general.

  18. Phospho-oligosaccharide dependent phosphorylation of ATP citrate lyase.

    PubMed

    Puerta, J; Mato, J M; Alemany, S

    1990-01-01

    The effect of insulin on ATP citrate lyase phosphorylation has been shown to be mimicked by a phospho-oligosaccharide in intact adipocytes. We demonstrate that the addition of phospho-oligosaccharide to intact adipocytes enhances the phosphorylation of ATP citrate lyase in the same tryptic peptide as insulin does. The addition of phospho-oligosaccharide to an adipocyte extract also results in an increase in ATP citrate lyase phosphorylation but in a different site than that observed in intact cells. The phospho-oligosaccharide-dependent incorporation of phosphate into ATP citrate lyase in intact cells is resistant to isopropanol and acetic acid, but the phosphoenzyme phosphorylated in cell extracts is acid labile. In cell extracts, the addition of phospho-oligosaccharide markedly inhibits ATP hydrolysis, which may explain the effect of this molecule on ATP citrate lyase phosphorylation in broken cells. These results support the hypothesis that this phospho-oligosaccharide mediates some of the effects of insulin on protein phosphorylation. They also indicate that caution should be exercised in interpreting the results obtained by adding phospho-oligosaccharide to broken cell preparations.

  19. Enzymatic Hydrolysis of Alginate to Produce Oligosaccharides by a New Purified Endo-Type Alginate Lyase

    PubMed Central

    Zhu, Benwei; Chen, Meijuan; Yin, Heng; Du, Yuguang; Ning, Limin

    2016-01-01

    Enzymatic hydrolysis of sodium alginate to produce alginate oligosaccharides has drawn increasing attention due to its advantages of containing a wild reaction condition, excellent gel properties and specific products easy for purification. However, the efficient commercial enzyme tools are rarely available. A new alginate lyase with high activity (24,038 U/mg) has been purified from a newly isolated marine strain, Cellulophaga sp. NJ-1. The enzyme was most active at 50 °C and pH 8.0 and maintained stability at a broad pH range (6.0–10.0) and temperature below 40 °C. It had broad substrate specificity toward sodium alginate, heteropolymeric MG blocks (polyMG), homopolymeric M blocks (polyM) and homopolymeric G blocks (polyG), and possessed higher affinity toward polyG (15.63 mM) as well as polyMG (23.90 mM) than polyM (53.61 mM) and sodium alginate (27.21 mM). The TLC and MS spectroscopy analysis of degradation products suggested that it completely hydrolyzed sodium alginate into oligosaccharides of low degrees of polymerization (DPs). The excellent properties would make it a promising tool for full use of sodium alginate to produce oligosaccharides. PMID:27275826

  20. Enzymatic Hydrolysis of Alginate to Produce Oligosaccharides by a New Purified Endo-Type Alginate Lyase.

    PubMed

    Zhu, Benwei; Chen, Meijuan; Yin, Heng; Du, Yuguang; Ning, Limin

    2016-06-06

    Enzymatic hydrolysis of sodium alginate to produce alginate oligosaccharides has drawn increasing attention due to its advantages of containing a wild reaction condition, excellent gel properties and specific products easy for purification. However, the efficient commercial enzyme tools are rarely available. A new alginate lyase with high activity (24,038 U/mg) has been purified from a newly isolated marine strain, Cellulophaga sp. NJ-1. The enzyme was most active at 50 °C and pH 8.0 and maintained stability at a broad pH range (6.0-10.0) and temperature below 40 °C. It had broad substrate specificity toward sodium alginate, heteropolymeric MG blocks (polyMG), homopolymeric M blocks (polyM) and homopolymeric G blocks (polyG), and possessed higher affinity toward polyG (15.63 mM) as well as polyMG (23.90 mM) than polyM (53.61 mM) and sodium alginate (27.21 mM). The TLC and MS spectroscopy analysis of degradation products suggested that it completely hydrolyzed sodium alginate into oligosaccharides of low degrees of polymerization (DPs). The excellent properties would make it a promising tool for full use of sodium alginate to produce oligosaccharides.

  1. Structural studies of sialylated oligosaccharides of human midcycle cervical mucin.

    PubMed

    Yurewicz, E C; Matsuura, F; Moghissi, K S

    1987-04-05

    It was previously shown that reductive alkali treatment of purified human cervical mucin releases a heterogeneous population of reduced neutral, sialylated, and sulfated oligosaccharides (Yurewicz, E. C., and Moghissi, K. S. (1981) J. Biol. Chem. 256, 11895-11904). Four major sialylated oligosaccharide fractions were isolated with approximate compositions of Fuc:GlcNac:Gal:NeuAc:N-acetylgalactosaminitol (GalNAcol) = 0:0:0:1:1 (B1a), 0:0:1:1:1 (B2b), 0:1:2:1:1 (B3a), and 1:1:2:1:1 (B4a), where Fuc is fucose. They comprised roughly 3, 11, 7, and 6% of recovered oligosaccharide chains, respectively. On the basis of periodate oxidations, methylation analyses, and sequential degradations with glycosidases, the following structures were determined. (Formula: see text) Oligosaccharides 1 and 2 are characterized by the presence of N-acetylneuraminic acid in alpha 2,6-linkage to N-acetylgalactosaminitol. The remaining oligosaccharides contain N-acetylneuraminic acid in alpha 2,3-linkage to galactose residues. Oligosaccharides 3 and 4 and oligosaccharides 5 and 6 were isolated as unresolved isomeric mixtures in fractions B3a and B4a, respectively. Oligosaccharides 3 and 4 were distinguished on the basis of susceptibility to digestion with Aspergillus niger beta-galactosidase whereas oligosaccharides 5 and 6 were distinguished on the basis of differential rates of digestion with beef kidney alpha-fucosidase. The structural data indicate the presence of at least two sialyltransferases in human cervical epithelium and further suggest a potential physiologically significant competition between sialyltransferase and beta-N-acetylglucosaminyltransferase for C-6 of the N-acetylgalactosamine residue O-glycosidically linked to serine/threonine of the polypeptide core.

  2. Combinational effects of prebiotic oligosaccharides on bifidobacterial growth and host gene expression in a simplified mixed culture model and neonatal mice.

    PubMed

    Ehara, Tatsuya; Izumi, Hirohisa; Tsuda, Muneya; Nakazato, Yuki; Iwamoto, Hiroshi; Namba, Kazuyoshi; Takeda, Yasuhiro

    2016-07-01

    It is important to provide formula-fed infants with a bifidobacteria-enriched gut microbiota similar to those of breastfed infants to ensure intestinal health. Prebiotics, such as certain oligosaccharides, are a useful solution to this problem, but the combinational benefits of these oligosaccharides have not been evaluated. This study investigated the benefits of oligosaccharide combinations and screened for an optimal combination of oligosaccharides to promote healthy gut microbiota of formula-fed infants. In vitro and in vivo experiments were performed to assess the bifidogenic effects of lactulose (LAC) alone and LAC combined with raffinose (RAF) and/or galacto-oligosaccharide (GOS), using a mixed culture model and neonatal mice orally administered with these oligosaccharides and Bifidobacterium breve. In the in vitro culture model, the combination of the three oligosaccharides (LAC-RAF-GOS) significantly increased cell numbers of B. breve and Bifidobacterium longum (P<0·05) compared with either LAC alone or the combination of two oligosaccharides, and resulted in the production of SCFA under anaerobic conditions. In the in vivo experiment, the LAC-RAF-GOS combination significantly increased cell numbers of B. breve and Bacteroidetes in the large intestinal content (P<0·05) and increased acetate concentrations in the caecal content and serum of neonatal mice. Genes related to metabolism and immune responses were differentially expressed in the liver and large intestine of mice administered with LAC-RAF-GOS. These results indicate a synergistic effect of the LAC-RAF-GOS combination on the growth of bifidobacteria and reveal possible benefits of this combination to the gut microbiota and health of infants.

  3. Resistant maltodextrin promotes fasting glucagon-like peptide-1 secretion and production together with glucose tolerance in rats.

    PubMed

    Hira, Tohru; Ikee, Asuka; Kishimoto, Yuka; Kanahori, Sumiko; Hara, Hiroshi

    2015-07-14

    Glucagon-like peptide-1 (GLP-1), which is produced and released from enteroendocrine L cells, plays pivotal roles in postprandial glycaemia. The ingestion of resistant maltodextrin (RMD), a water-soluble non-digestible saccharide, improves the glycaemic response. In the present study, we examined whether the continuous feeding of RMD to rats affected GLP-1 levels and glycaemic control. Male Sprague-Dawley rats (6 weeks of age) were fed an American Institute of Nutrition (AIN)-93G-based diet containing either cellulose (5 %) as a control, RMD (2.5 or 5 %), or fructo-oligosaccharides (FOS, 2.5 or 5 %) for 7 weeks. During the test period, an intraperitoneal glucose tolerance test (IPGTT) was performed after 6 weeks. Fasting GLP-1 levels were significantly higher in the 5 % RMD group than in the control group after 6 weeks. The IPGTT results showed that the glycaemic response was lower in the 5 % RMD group than in the control group. Lower caecal pH, higher caecal tissue and content weights were observed in the RMD and FOS groups. Proglucagon mRNA levels were increased in the caecum and colon of both RMD and FOS groups, whereas caecal GLP-1 content was increased in the 5 % RMD group. In addition, a 1 h RMD exposure induced GLP-1 secretion in an enteroendocrine L-cell model, and single oral administration of RMD increased plasma GLP-1 levels in conscious rats. The present study demonstrates that continuous ingestion of RMD increased GLP-1 secretion and production in normal rats, which could be stimulated by its direct and indirect (enhanced gut fermentation) effects on GLP-1-producing cells, and contribute to improving glucose tolerance.

  4. Modification of linear (β1→3)-linked gluco-oligosaccharides with a novel recombinant β-glucosyltransferase (trans-β-glucosidase) enzyme from Bradyrhizobium diazoefficiens.

    PubMed

    Dobruchowska, Justyna M; Jonsson, Jon O; Fridjonsson, Olafur H; Aevarsson, Arnthor; Kristjansson, Jakob K; Altenbuchner, Josef; Watzlawick, Hildegard; Gerwig, Gerrit J; Dijkhuizen, Lubbert; Kamerling, Johannis P; Hreggvidsson, Gudmundur O

    2016-11-01

    Recently, we have shown that glycoside hydrolases enzymes of family GH17 from proteobacteria (genera Pseudomonas, Azotobacter) catalyze elongation transfer reactions with laminari-oligosaccharides generating (β1→3) linkages preferably and to a lesser extent (β1→6) or (β1→4) linkages. In the present study, the cloning and characterization of the gene encoding the structurally very similar GH17 domain of the NdvB enzyme from Bradyrhizobium diazoefficiens, designated Glt20, as well as its catalytic properties are described. The Glt20 enzyme was strikingly different from the previously investigated bacterial GH17 enzymes, both regarding substrate specificity and product formation. The Azotobacter and Pseudomonas enzymes cleaved the donor laminari-oligosaccharide substrates three or four moieties from the non-reducing end, generating linear oligosaccharides. In contrast, the Glt20 enzyme cleaved donor laminari-oligosaccharide substrates two glucose moieties from the reducing end, releasing laminaribiose and transferring the remainder to laminari-oligosaccharide acceptor substrates creating only (β1→3)(β1→6) branching points. This enables Glt20 to transfer larger oligosaccharide chains than the other type of bacterial enzymes previously described, and helps explain the biologically significant formation of cyclic β-glucans in B. diazoefficiens. © The Author 2016. Published by Oxford University Press. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  5. Structural elucidation of an asparagine-linked oligosaccharide from the hyperthermophilic archaeon, Archaeoglobus fulgidus.

    PubMed

    Fujinami, Daisuke; Nyirenda, James; Matsumoto, Shunsuke; Kohda, Daisuke

    2015-09-02

    The genome of the hyperthermophilic archaeon, Archaeoglobus fulgidus, contains three paralogous AglB genes that encode oligosaccharyltransferase (OST) proteins. The OST enzymes catalyze the transfer of an oligosaccharide chain from lipid-linked oligosaccharides (LLO) to asparagine residues in proteins. The detergent-solubilized membrane fractions prepared from cultured A. fulgidus cells contain both OST and LLO. The addition of a peptide containing the glycosylation sequon produced oligosaccharide chains attached to a structurally defined peptide. To facilitate the NMR analysis, the cells were grown in rich medium supplemented with (13)C-glucose, to label the LLOs metabolically. The MS analysis of the glycopeptide revealed that the glucose and galactose residues were nearly fully (13)C-labeled, but the mannose residues were fractionally labeled with about 20% efficiency. An immunodetection experiment revealed that the longest AglB paralog (AfAglB-L) was expressed in the membrane fractions under our cell culture conditions, while the other two shorter AglB paralogs (AfAglB-S1 and AfAglB-S2) were not. Thus, the oligosaccharide chain analyzed in this study was the product of AfAglB-L. The N-glycan consists of eight hexose residues, as follows: The α1,3-linked glucose is an optional residue branching from the distal mannose residue. The MS analysis of the minor HPLC peak of the in vitro oligosaccharyl transfer products also revealed an optional sulfate modification on the glucose residue directly linked to the Asn residue. The present data will be useful for structural and functional studies of the N-glycosylation system of A. fulgidus.

  6. Branching N-linked oligosaccharides in breast cancer.

    PubMed

    Korczak, B; Goss, P; Fernandez, B; Baker, M; Dennis, J W

    1994-01-01

    Tumor progression in rodent and human tumors is commonly associated with changes in glycoprotein glycosylation, in particular increased beta 1-6GlcNAc-branching, a regulatory step in expression of polylactosamine and extended-chain Lewis antigens. Loss of the branched oligosaccharides in murine tumor cells either due to somatic mutation, or treatment of the cells with the oligosaccharide processing inhibitor swainsonine, blocks tumor cells invasion in vitro and reduces solid tumor growth in vivo. Swainsonine and other inhibitors of N-linked oligosaccharide processing may be useful anti-cancer drugs, a premise which has begun to be tested in humans.

  7. High mannose oligosaccharide of phytohemagglutinin is attached to asparagine 12 and the modified oligosaccharide to asparagine 60. [Phaseolus vulgaris

    SciTech Connect

    Sturm, A.; Chrispeels, M.J.

    1986-05-01

    Phytohemagglutinin, the lectin of the common bean Phaseolus vulgaris, has a high mannose and a modified (fucosylated) oligosaccharide on each polypeptide. Fractionation by high performance liquid chromatography of tryptic digests of (/sup 3/H)fucose or (/sup 3/H)glucosamine labeled phytohemagglutinin, followed by amino acid sequencing of the isolated glycopeptides, shows that the high mannose oligosaccharide is attached to Asn/sup 12/ and the modified oligosaccharide to Asn /sup 60/ of the protein. In animal glycoproteins, high mannose chains are rarely found at the N-terminal side of complex chains.

  8. Determining the isomeric heterogeneity of neutral oligosaccharide-alditols of bovine submaxillary mucin using negative ion traveling wave ion mobility mass spectrometry.

    PubMed

    Li, Hongli; Bendiak, Brad; Siems, William F; Gang, David R; Hill, Herbert H

    2015-02-17

    Negative ions produced by electrospray ionization were used to evaluate the isomeric heterogeneity of neutral oligosaccharide-alditols isolated from bovine submaxillary mucin (BSM). The oligosaccharide-alditol mixture was preseparated on an off-line high-performance liquid chromatography (HPLC) column, and the structural homogeneity of individual LC fractions was investigated using a Synapt G2 traveling wave ion mobility spectrometer coupled between quadupole and time-of-flight mass spectrometers. Mixtures of isomers separated by both chromatography and ion mobility spectrometry were studied. Tandem mass spectrometry (MS/MS) of multiple mobility peaks having the same mass-to-charge ratio (m/z) demonstrated the presence of different structural isomers and not differences in ion conformations due to charge site location. Although the oligosaccharide-alditol mixture was originally separated by HPLC, multiple ion mobility peaks due to structural isomers were observed for a number of oligosaccharide-alditols from single LC fractions. The collision-induced dissociation cells located in front of and after the ion mobility separation device enabled oligosaccharide precursor or product ions to be separated by ion mobility and independent fragmentation spectra to be acquired for isomeric carbohydrate precursor or product ions. MS/MS spectra so obtained for independent mobility peaks at a single m/z demonstrated the presence of structural variants or stereochemical isomers having the same molecular formula. This was observed both for oligosaccharide precursor and product ions. In addition, mobilities of both [M - H](-) and [M + Cl](-) ions, formed by adding NH4OH or NH4Cl to the electrospray solvent, were examined and compared for selected oligosaccharide-alditols. Better separation among structural isomers appeared to be achieved for some [M + Cl](-) anions.

  9. Multiple applications of ion chromatography oligosaccharide fingerprint profiles to solve a variety of sugar and sugar-biofuel industry problems.

    PubMed

    Eggleston, Gillian; Borges, Eduardo

    2015-03-25

    Sugar crops contain a broad variety of carbohydrates used for human consumption and the production of biofuels and bioproducts. Ion chromatography with integrated pulsed amperometric detection (IC-IPAD) can be used to simultaneously detect mono-, di-, and oligosaccharides, oligosaccharide isomers, mannitol, and ethanol in complex matrices from sugar crops. By utilizing a strong NaOH/NaOAc gradient method over 45 min, oligosaccharides of at least 2-12 dp can be detected. Fingerprint IC oligosaccharide profiles are extremely selective, sensitive, and reliable and can detect deterioration product metabolites from as low as 100 colony-forming units/mL lactic acid bacteria. The IC fingerprints can also be used to (i) monitor freeze deterioration, (ii) optimize harvesting methods and cut-to-crush times, (iii) differentiate between white refined sugar from sugar cane and from sugar beets, (iv) verify the activities of carbohydrate enzymes, (v) select yeasts for ethanol fermentations, and (vi) isolate and diagnose infections and processing problems in sugar factories.

  10. Acetylated Chitosan Oligosaccharides Act as Antagonists against Glutamate-Induced PC12 Cell Death via Bcl-2/Bax Signal Pathway

    PubMed Central

    Hao, Cui; Gao, Lixia; Zhang, Yiran; Wang, Wei; Yu, Guangli; Guan, Huashi; Zhang, Lijuan; Li, Chunxia

    2015-01-01

    Chitosan oligosaccharides (COSs), depolymerized products of chitosan composed of β-(1→4) d-glucosamine units, have broad range of biological activities such as antitumour, antifungal, and antioxidant activities. In this study, peracetylated chitosan oligosaccharides (PACOs) and N-acetylated chitosan oligosaccharides (NACOs) were prepared from the COSs by chemcal modification. The structures of these monomers were identified using NMR and ESI-MS spectra. Their antagonist effects against glutamate-induced PC12 cell death were investigated. The results showed that pretreatment of PC12 cells with the PACOs markedly inhibited glutamate-induced cell death in a concentration-dependent manner. The PACOs were better glutamate antagonists compared to the COSs and the NACOs, suggesting the peracetylation is essential for the neuroprotective effects of chitosan oligosaccharides. In addition, the PACOs pretreatment significantly reduced lactate dehydrogenase release and reactive oxygen species production. It also attenuated the loss of mitochondrial membrane potential. Further studies indicated that the PACOs inhibited glutamate-induced cell death by preventing apoptosis through depressing the elevation of Bax/Bcl-2 ratio and caspase-3 activation. These results suggest that PACOs might be promising antagonists against glutamate-induced neural cell death. PMID:25775423

  11. Stable isotope labeling of oligosaccharide cell surface antigens

    SciTech Connect

    Unkefer, C.J.; Silks, L.A. III; Martinez, R.A.

    1998-12-31

    The overall goal of this Laboratory Directed Research and Development (LDRD) project was to develop new methods for synthesis of {sup 13}C-labeled oligosaccharides that are required for nuclear magnetic resonance (NMR) studies of their solution conformation. Oligosaccharides are components of the cell`s outer surface and are involved in important processes such as cell-cell recognition and adhesion. Recently, Danishefsky and coworkers at Slone-Kettering Cancer Center developed a method for the solid-phase chemical synthesis of oligosaccharides. The specific goal of this LDRD project was to prepare uniform {sup 13}C-labeled aldohexose precursors required for the solid-phase synthesis of the Lewis blood-group antigenic determinants. We report the synthesis of {sup 13}C-labeled D-glucal, D-galactal and Fucosyl precursors. We have been collaborating with the Danishefsky group on the synthesis of the Lewis oligosaccharides and the NMR analysis of their solution conformation.

  12. Applications of Mass Spectrometry to Structural Analysis of Marine Oligosaccharides

    PubMed Central

    Lang, Yinzhi; Zhao, Xia; Liu, Lili; Yu, Guangli

    2014-01-01

    Marine oligosaccharides have attracted increasing attention recently in developing potential drugs and biomaterials for their particular physical and chemical properties. However, the composition and sequence analysis of marine oligosaccharides are very challenging for their structural complexity and heterogeneity. Mass spectrometry (MS) has become an important technique for carbohydrate analysis by providing more detailed structural information, including molecular mass, sugar constituent, sequence, inter-residue linkage position and substitution pattern. This paper provides an overview of the structural analysis based on MS approaches in marine oligosaccharides, which are derived from some biologically important marine polysaccharides, including agaran, carrageenan, alginate, sulfated fucan, chitosan, glycosaminoglycan (GAG) and GAG-like polysaccharides. Applications of electrospray ionization mass spectrometry (ESI-MS) are mainly presented and the general applications of matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) are also outlined. Some technical challenges in the structural analysis of marine oligosaccharides by MS have also been pointed out. PMID:24983643

  13. Applications of mass spectrometry to structural analysis of marine oligosaccharides.

    PubMed

    Lang, Yinzhi; Zhao, Xia; Liu, Lili; Yu, Guangli

    2014-06-30

    Marine oligosaccharides have attracted increasing attention recently in developing potential drugs and biomaterials for their particular physical and chemical properties. However, the composition and sequence analysis of marine oligosaccharides are very challenging for their structural complexity and heterogeneity. Mass spectrometry (MS) has become an important technique for carbohydrate analysis by providing more detailed structural information, including molecular mass, sugar constituent, sequence, inter-residue linkage position and substitution pattern. This paper provides an overview of the structural analysis based on MS approaches in marine oligosaccharides, which are derived from some biologically important marine polysaccharides, including agaran, carrageenan, alginate, sulfated fucan, chitosan, glycosaminoglycan (GAG) and GAG-like polysaccharides. Applications of electrospray ionization mass spectrometry (ESI-MS) are mainly presented and the general applications of matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) are also outlined. Some technical challenges in the structural analysis of marine oligosaccharides by MS have also been pointed out.

  14. Synbiotic matrices derived from plant oligosaccharides and polysaccharides

    USDA-ARS?s Scientific Manuscript database

    A porous synbiotic matrix was prepared by lyophilization of alginate and pectin or fructan oligosaccharides and polysaccharides cross-linked with calcium. These synbiotic matrices were excellent physical structures to support the growth of Lactobacillus acidophilus (1426) and Lactobacillus reuteri (...

  15. An Overview of the Protective Effects of Chitosan and Acetylated Chitosan Oligosaccharides against Neuronal Disorders.

    PubMed

    Hao, Cui; Wang, Wei; Wang, Shuyao; Zhang, Lijuan; Guo, Yunliang

    2017-03-23

    Chitin is the second most abundant biopolymer on Earth and is mainly comprised of a marine invertebrate, consisting of repeating β-1,4 linked N-acetylated glucosamine units, whereas its N-deacetylated product, chitosan, has broad medical applications. Interestingly, chitosan oligosaccharides have therapeutic effects on different types of neuronal disorders, including, but not limited to, Alzheimer's disease, Parkinson's disease, and nerve crush injury. A common link among neuronal disorders is observed at a sub-cellular level, such as atypical protein assemblies and induced neuronal death. Chronic activation of innate immune responses that lead to neuronal injury is also common in these diseases. Thus, the common mechanisms of neuronal disorders might explain the general therapeutic effects of chitosan oligosaccharides and their derivatives in these diseases. This review provides an update on the pathogenesis and therapy for neuronal disorders and will be mainly focused on the recent progress made towards the neuroprotective properties of chitosan and acetylated chitosan oligosaccharides. Their structural features and the underlying molecular mechanisms will also be discussed.

  16. In vitro investigation into the potential prebiotic activity of honey oligosaccharides.

    PubMed

    Sanz, María Luz; Polemis, Nikolaos; Morales, Valle; Corzo, Nieves; Drakoularakou, Alexandra; Gibson, Glenn R; Rastall, Robert A

    2005-04-20

    The effect of honey oligosaccharides on the growth of fecal bacteria was studied using an in vitro fermentation system. Prior to treatment, glucose and fructose (31.73 and 21.41 g/100 g of product, respectively) present in honey, which would be digested in the upper gut, were removed to avoid any influence on bacterial populations in the fermentations. Nanofiltration, yeast (Saccharomyces cerevisiae) treatment, and adsorption onto activated charcoal were used to remove monosaccharides. Prebiotic (microbial fermentation) activities of the three honey oligosaccharide fractions and the honey sample were studied and compared with fructooligosaccharide (FOS), using 1% (w/v) fecal bacteria in an in vitro fermentation system (10 mg of carbohydrate, 1.0 mL of basal medium). A prebiotic index (PI) was calculated for each carbohydrate source. Honey oligosaccharides seem to present potential prebiotic activity (PI values between 3.38 and 4.24), increasing the populations of bifidobacteria and lactobacilli, although not to the levels of FOS (PI of 6.89).

  17. Milk Proteins, Peptides, and Oligosaccharides: Effects against the 21st Century Disorders

    PubMed Central

    Hsieh, Chia-Chien; Hernández-Ledesma, Blanca; Fernández-Tomé, Samuel; Weinborn, Valerie; Barile, Daniela; de Moura Bell, Juliana María Leite Nobrega

    2015-01-01

    Milk is the most complete food for mammals, as it supplies all the energy and nutrients needed for the proper growth and development of the neonate. Milk is a source of many bioactive components, which not only help meeting the nutritional requirements of the consumers, but also play a relevant role in preventing various disorders. Milk-derived proteins and peptides have the potential to act as coadjuvants in conventional therapies, addressing cardiovascular diseases, metabolic disorders, intestinal health, and chemopreventive properties. In addition to being a source of proteins and peptides, milk contains complex oligosaccharides that possess important functions related to the newborn's development and health. Some of the health benefits attributed to milk oligosaccharides include prebiotic probifidogenic effects, antiadherence of pathogenic bacteria, and immunomodulation. This review focuses on recent findings demonstrating the biological activities of milk peptides, proteins, and oligosaccharides towards the prevention of diseases of the 21st century. Processing challenges hindering large-scale production and commercialization of those bioactive compounds have been also addressed. PMID:25789308

  18. Major carbohydrate, polyol, and oligosaccharide profiles of agave syrup. Application of this data to authenticity analysis.

    PubMed

    Willems, Jamie L; Low, Nicholas H

    2012-09-05

    Nineteen pure agave syrups representing the three major production regions and four processing facilities in Mexico were analyzed for their major carbohydrate, polyol, and oligosaccharide profiles, as well as their physicochemical properties (pH, °Brix, total acidity, percent total titratable acidity, and color). Additionally, the detection of intentional debasing of agave syrup with four commercial nutritive sweeteners (HFCS 55 and 90, DE 42 and sucrose) was afforded by oligosaccharide profiling employing both high performance anion exchange liquid chromatography with pulsed amperometric detection (HPAE-PAD) and capillary gas chromatography with flame ionization detection (CGC-FID). Results showed that the major carbohydrate and polyol in agave syrups were fructose and inositol with mean concentrations of 84.29% and 0.38%, respectively. Oligosaccharide profiling was extremely successful for adulteration detection with detection limits ranging from 0.5 to 2.0% for the aforementioned debasing agents. Also, all four of these possible adulterants could be detected within a single chromatographic analysis.

  19. Milk Oligosaccharides Inhibit Human Rotavirus Infectivity in MA104 Cells.

    PubMed

    Laucirica, Daniel R; Triantis, Vassilis; Schoemaker, Ruud; Estes, Mary K; Ramani, Sasirekha

    2017-09-01

    Background: Oligosaccharides in milk act as soluble decoy receptors and prevent pathogen adhesion to the infant gut. Milk oligosaccharides reduce infectivity of a porcine rotavirus strain; however, the effects on human rotaviruses are less well understood.Objective: In this study, we determined the effect of specific and abundant milk oligosaccharides on the infectivity of 2 globally dominant human rotavirus strains.Methods: Four milk oligosaccharides-2'-fucosyllactose (2'FL), 3'-sialyllactose (3'SL), 6'-sialyllactose (6'SL), and galacto-oligosaccharides-were tested for their effects on the infectivity of human rotaviruses G1P[8] and G2P[4] through fluorescent focus assays on African green monkey kidney epithelial cells (MA104 cells). Oligosaccharides were added at different time points in the infectivity assays. Infections in the absence of oligosaccharides served as controls.Results: When compared with infections in the absence of glycans, all oligosaccharides substantially reduced the infectivity of both human rotavirus strains in vitro; however, virus strain-specific differences in effects were observed. Compared with control infections, the maximum reduction in G1P[8] infectivity was seen with 2'FL when added after the onset of infection (62% reduction, P < 0.01), whereas the maximum reduction in G2P[4] infectivity was seen with the mixture of 3'SL + 6'SL when added during infection (73% reduction, P < 0.01). The mixture of 3'SL + 6'SL at the same ratio as is present in breast milk was more potent in reducing G2P[4] infectivity (73% reduction, P < 0.01) than when compared with 3'SL (47% reduction) or 6'SL (40% reduction) individually. For all oligosaccharides the reduction in infectivity was mediated by an effect on the virus and not on the cells.Conclusions: Milk oligosaccharides reduce the infectivity of human rotaviruses in MA104 cells, primarily through an effect on the virus. Although breastfed infants are directly protected, the addition of specific

  20. Recent Advances in the Synthesis of C-oligosaccharides

    PubMed Central

    Yuan, Xuejun; Linhardt, Robert J.

    2014-01-01

    This paper reviews the recent advances in the synthesis of catabolically stable sugar mimetics, C-oligosaccharides. These compounds are synthetic analogs of the naturally occurring O-oligosaccharides, in which the interglycosidic oxygen has been replaced by a methylene group. This review is organized in terms of chemistry used to assemble C-oligosaccarides under the sub-headings: anionic approaches, cationic methods, reductive glycosyl samarium chemistry, cyclization methodology, and free radical chemistry. PMID:16305537

  1. Identification of Oligosaccharides in Feces of Breast-fed Infants and Their Correlation with the Gut Microbial Community.

    PubMed

    Davis, Jasmine C C; Totten, Sarah M; Huang, Julie O; Nagshbandi, Sadaf; Kirmiz, Nina; Garrido, Daniel A; Lewis, Zachery T; Wu, Lauren D; Smilowitz, Jennifer T; German, J Bruce; Mills, David A; Lebrilla, Carlito B

    2016-09-01

    Glycans in breast milk are abundant and found as either free oligosaccharides or conjugated to proteins and lipids. Free human milk oligosaccharides (HMOs) function as prebiotics by stimulating the growth of beneficial bacteria while preventing the binding of harmful bacteria to intestinal epithelial cells. Bacteria have adapted to the glycan-rich environment of the gut by developing enzymes that catabolize glycans. The decrease in HMOs and the increase in glycan digestion products give indications of the active enzymes in the microbial population. In this study, we quantitated the disappearance of intact HMOs and characterized the glycan digestion products in the gut that are produced by the action of microbial enzymes on HMOs and glycoconjugates from breast milk. Oligosaccharides from fecal samples of exclusively breast-fed infants were extracted and profiled using nanoLC-MS. Intact HMOs were found in the fecal samples, additionally, other oligosaccharides were found corresponding to degraded HMOs and non-HMO based compounds. The latter compounds were fragments of N-glycans released through the cleavage of the linkage to the asparagine residue and through cleavage of the chitobiose core of the N-glycan. Marker gene sequencing of the fecal samples revealed bifidobacteria as the dominant inhabitants of the infant gastrointestinal tracts. A glycosidase from Bifidobacterium longum subsp. longum was then expressed to digest HMOs in vitro, which showed that the digested oligosaccharides in feces corresponded to the action of glycosidases on HMOs. Similar expression of endoglycosidases also showed that N-glycans were released by bacterial enzymes. Although bifidobacteria may dominate the gut, it is possible that specific minority species are also responsible for the major products observed in feces. Nonetheless, the enzymatic activity correlated well with the known glycosidases in the respective bacteria, suggesting a direct relationship between microbial abundances and

  2. Beneficial effects of human milk oligosaccharides on gut microbiota.

    PubMed

    Musilova, S; Rada, V; Vlkova, E; Bunesova, V

    2014-09-01

    Human milk is the gold standard for nourishment of early infants because it contains a number of bioactive components, such as human milk oligosaccharides (HMOs). The high concentration and structural diversity of HMOs are unique to humans. HMOs are a group of complex and diverse glycans that are resistant to gastrointestinal digestion and reach the infant colon as the first prebiotics. N-acetyl-glucosamine containing oligosaccharides were first identified 50 years ago as the 'bifidus factor', a selective growth substrate for intestinal bifidobacteria, thus providing a conceptual basis for HMO-specific bifidogenic activity. Bifidobacterial species are the main utilisers of HMOs in the gastrointestinal tract and represent the dominant microbiota of breast-fed infants, and they may play an important role in maintaining the general health of newborn children. Oligosaccharides are also known to directly interact with the surface of pathogenic bacteria, and various oligosaccharides in milk are believed to inhibit the binding of pathogens and toxins to host cell receptors. Furthermore, HMOs are thought to contribute to the development of infant intestine and brain. Oligosaccharides currently added to infant formula are structurally different from the oligosaccharides naturally occurring in human milk and, therefore, they are unlikely to mimic some of the structure-specific effects. In this review, we describe how HMOs can modulate gut microbiota. This article summarises information up to date about the relationship between the intestinal microbiota and HMOs, and other possible indirect effects of HMOs on intestinal environment.

  3. [Structural and functional aspects of oligosaccharides in human milk].

    PubMed

    Kunz, C; Rudloff, S

    1996-03-01

    About a century ago, pediatricians observed that in feces of breast-fed infants, compared to those of bottle-fed infants, Bifidobacterium bifidum was the predominant microorganism. It was shown thereafter that aminosugar-containing oligosaccharides are growth factors for a specific strain of Bifidobacterium. Meanwhile, more than 130 lactose-derived oligosaccharides have been identified in human milk. Some of these oligosaccharides like Lacto-N-Tetraose and Lacto-N-Fucopentaose I and II do not occur in minute amounts but in concentrations up to 1-2 g/L. As the total amount of complex oligosaccharides is between 3-6 g/L those components have to be considered as major human milk constituents. There is striking evidence that human milk oligosaccharides are potent inhibitors of bacterial adhesion to epithelial surfaces, an initial stage of infective processes. Therefore, these oligosaccharides are considered to be soluble receptor analogues of epithelial cell surfaces participating in the non-immunological defense system of human milk-fed infants.

  4. Chemical Characterization of Potentially Prebiotic Oligosaccharides in Brewed Coffee and Spent Coffee Grounds.

    PubMed

    Tian, Tian; Freeman, Samara; Corey, Mark; German, J Bruce; Barile, Daniela

    2017-04-05

    Oligosaccharides are indigestible carbohydrates widely present in mammalian milk and in some plants. Milk oligosaccharides are associated with positive health outcomes; however, oligosaccharides in coffee have not been extensively studied. We investigated the oligosaccharides and their monomeric composition in dark roasted coffee beans, brewed coffee, and spent coffee grounds. Oligosaccharides with a degree of polymerization ranging from 3 to 15, and their constituent monosaccharides, were characterized and quantified. The oligosaccharides identified were mainly hexoses (potentially galacto-oligosaccharides and manno-oligosaccharides) containing a heterogeneous mixture of glucose, arabinose, xylose, and rhamnose. The diversity of oligosaccharides composition found in these coffee samples suggests that they could have selective prebiotic activity toward specific bacterial strains able to deconstruct the glycosidic bonds and utilize them as a carbon source.

  5. A computer-controlled system to simulate conditions of the large intestine with peristaltic mixing, water absorption and absorption of fermentation products.

    PubMed

    Minekus, M; Smeets-Peeters, M; Bernalier, A; Marol-Bonnin, S; Havenaar, R; Marteau, P; Alric, M; Fonty, G; Huis in't Veld, J H

    1999-12-01

    This paper introduces a new type of system to simulate conditions in the large intestine. This system combines removal of metabolites and water with peristaltic mixing to obtain and handle physiological concentrations of microorganisms, dry matter and microbial metabolites. The system has been designed to be complementary to the dynamic multi-compartmental system that simulates conditions in the stomach and small intestine described by Minekus et al. [Minekus M, Marteau P, Havenaar R, Huis in't Veld JHJ (1995) ATLA 23:197-209]. High densities of microorganisms, comparable to those found in the colon in vivo, were achieved by absorption of water and dialysis of metabolites through hollow-fibre membranes inside the reactor compartments. The dense chyme was mixed and transported by peristaltic movements. The potential of the system as a tool to study fermentation was demonstrated in experiments with pectin, fructo-oligosaccharide, lactulose and lactitol as substrates. Parameters such as total acid production and short-chain fatty acid (SCFA) patterns were determined with time to characterize the fermentation. The stability of the microflora in the system was tested after inoculation with fresh fecal samples and after inoculation with a microflora that was maintained in a fermenter. Both approaches resulted in total anaerobic bacterial counts higher than 10(10) colony-forming units/ml with physiological levels of Bifidobacterium, Lactobacillus, Enterobacteriaceae and Clostridium. The dry matter content was approximately 10%, while the total SCFA concentration was maintained at physiological concentrations with similar molar ratios for acetic acid, propionic acid and butyric acid as measured in vivo.

  6. Typing of blood-group antigens on neutral oligosaccharides by negative-ion electrospray ionization tandem mass spectrometry.

    PubMed

    Zhang, Hongtao; Zhang, Shuang; Tao, Guanjun; Zhang, Yibing; Mulloy, Barbara; Zhan, Xiaobei; Chai, Wengang

    2013-06-18

    Blood-group antigens, such as those containing fucose and bearing the ABO(H)- and Lewis-type determinants expressed on the carbohydrate chains of glycoproteins and glycolipids, and also on unconjugated free oligosaccharides in human milk and other secretions, are associated with various biological functions. We have previously shown the utility of negative-ion electrospay ionization tandem mass spectrometry with collision-induced dissociation (ESI-CID-MS/MS) for typing of Lewis (Le) determinants, for example, Le(a), Le(x), Le(b), and Le(y) on neutral and sialylated oligosaccharide chains. In the present report, we extended the strategy to characterization of blood-group A-, B-, and H-determinants on type 1 and type 2 and also on type 4 globoside chains to provide a high sensitivity method for typing of all the major blood-group antigens, including the A, B, H, Le(a), Le(x), Le(b), and Le(y) determinants, present in oligosaccharides. Using the principles established, we identified two minor unknown oligosaccharide components present in the products of enzymatic synthesis by bacterial fermentation. We also demonstrated that the unique fragmentations derived from the D- and (0,2)A-type cleavages observed in ESI-CID-MS/MS, which are important for assigning blood-group and chain types, only occur under the negative-ion conditions for reducing sugars but not for reduced alditols or under positive-ion conditions.

  7. Comparison of the cryoprotective effects of trehalose, alginate, and its oligosaccharides on peeled shrimp (Litopenaeus vannamei) during frozen storage.

    PubMed

    Ma, Lu-kai; Zhang, Bin; Deng, Shang-gui; Xie, Chao

    2015-03-01

    The cryoprotective effects of trehalose, alginate, and its oligosaccharides on peeled shrimp (Litopenaeus vannamei) during frozen storage was investigated by monitoring thawing loss, color, texture, myofibrillar protein content, Ca2+ -ATPase activity, and performing microscopic structural analysis. Data revealed significant (p < 0.05) inhibitory effects on thawing loss and textural variables (springiness and chewiness) in trehalose-, alginate oligosaccharides-, and sodium pyrophosphate-treated shrimp compared with the control and alginate-treated batches. L* values revealed that these saccharides had a positive effect on color stability during frozen storage. In addition, the results of chemical analyses showed that trehalose and alginate oligosaccharide treatments effectively maintained an increased myofibrillar protein content and Ca2+ -ATPase activity in frozen shrimp. In addition, hematoxylin & eosin staining and SDS-PAGE confirmed that these cryoprotective saccharides slowed the degradation of muscle proteins and the damage to muscle tissue structures. Overall, the application of trehalose and alginate oligosaccharides to peeled frozen shrimp might maintain better quality and extend the commercialization of these refrigerated products.

  8. Isolation and Structural Characterization of an Oligosaccharide Produced by Bacillus subtilis in a Maltose-Containing Medium

    PubMed Central

    Shin, Kwang-Soon

    2016-01-01

    Among 116 bacterial strains isolated from Korean fermented foods, one strain (SS-76) was selected for producing new oligosaccharides in a basal medium containing maltose as the sole source of carbon. Upon morphological characterization using scanning electron microscopy, the cells of strain SS-76 appeared rod-shaped; subsequent 16S rRNA gene sequence analysis revealed that strain SS-76 was phylogenetically close to Bacillus subtilis. The main oligosaccharide fraction B extracted from the culture supernatant of B. subtilis SS-76 was purified by high performance liquid chromatography. Subsequent structural analysis revealed that this oligosaccharide consisted only of glucose, and methylation analysis indicated similar proportions of glucopyranosides in the 6-linkage, 4-linkage, and non-reducing terminal positions. Matrix-assisted laser-induced/ionization time-of-flight/mass spectrometry and electrospray ionization-based liquid chromatography-mass spectrometry/mass spectrometry analyses suggested that this oligosaccharide consisted of a trisaccharide unit with 1,6- and 1,4-glycosidic linkages. The anomeric signals in the 1H-nuclear magnetic resonance spectrum corresponded to α-anomeric configurations, and the trisaccharide was finally identified as panose (α-D-glucopyranosyl-1,6-α-D-glucopyranosyl-1,4-D-glucose). These results suggest that B. subtilis SS-76 converts maltose into panose; strain SS-76 may thus find industrial application in the production of panose. PMID:27390729

  9. Isolation and Structural Characterization of an Oligosaccharide Produced by Bacillus subtilis in a Maltose-Containing Medium.

    PubMed

    Shin, Kwang-Soon

    2016-06-01

    Among 116 bacterial strains isolated from Korean fermented foods, one strain (SS-76) was selected for producing new oligosaccharides in a basal medium containing maltose as the sole source of carbon. Upon morphological characterization using scanning electron microscopy, the cells of strain SS-76 appeared rod-shaped; subsequent 16S rRNA gene sequence analysis revealed that strain SS-76 was phylogenetically close to Bacillus subtilis. The main oligosaccharide fraction B extracted from the culture supernatant of B. subtilis SS-76 was purified by high performance liquid chromatography. Subsequent structural analysis revealed that this oligosaccharide consisted only of glucose, and methylation analysis indicated similar proportions of glucopyranosides in the 6-linkage, 4-linkage, and non-reducing terminal positions. Matrix-assisted laser-induced/ionization time-of-flight/mass spectrometry and electrospray ionization-based liquid chromatography-mass spectrometry/mass spectrometry analyses suggested that this oligosaccharide consisted of a trisaccharide unit with 1,6- and 1,4-glycosidic linkages. The anomeric signals in the (1)H-nuclear magnetic resonance spectrum corresponded to α-anomeric configurations, and the trisaccharide was finally identified as panose (α-D-glucopyranosyl-1,6-α-D-glucopyranosyl-1,4-D-glucose). These results suggest that B. subtilis SS-76 converts maltose into panose; strain SS-76 may thus find industrial application in the production of panose.

  10. Competitive inhibition of cellobiohydrolase I by manno-oligosaccharides.

    PubMed

    Xin, Donglin; Ge, Xiaoyan; Sun, Zongping; Viikari, Liisa; Zhang, Junhua

    2015-01-01

    In the hydrolysis of softwood, significant amounts of manno-oligosaccharides (MOS) are released from mannan, the major hemicelluloses in softwood. However, the impact of MOS on the performance of cellulases is not yet clear. In this work, the effect of mannan and MOS in cellulose hydrolysis by cellulases, especially cellobiohydrolase I (CBHI) from Thermoascus aurantiacus (Ta Cel7A), was studied. The glucose yield of Avicel decreased with an increasing amount of added mannan. Commercial cellulases contained mannan hydrolysing enzymes, and β-glucosidase played an important role in mannan hydrolysis. Addition of 10mg/ml mannan reduced the glucose yield of Avicel (at 20g/l) from 40.1 to 24.3%. No inhibition of β-glucosidase by mannan was observed. The negative effects of mannan and MOS on the hydrolytic action of cellulases indicated that the inhibitory effect was at least partly attributed to the inhibition of Ta Cel7A (CBHI), but not on β-glucosidase. Kinetic experiments showed that MOS were competitive inhibitors of the CBHI from T. aurantiacus, and mannobiose had a stronger inhibitory effect on CBHI than mannotriose or mannotetraose. For efficient hydrolysis of softwood, it was necessary to add supplementary enzymes to hydrolyze both mannan and MOS to less inhibitory product, mannose. Copyright © 2014 Elsevier Inc. All rights reserved.

  11. Thermophoresis of cyclic oligosaccharides in polar solvents.

    PubMed

    Eguchi, Kazuya; Niether, Doreen; Wiegand, Simone; Kita, Rio

    2016-09-01

    Cyclodextrins are cyclic oligosaccharides which are interesting as drug delivery systems, because they can be used as containers for pharmaceutical substances. We studied the Ludwig-Soret effect of [Formula: see text]-, [Formula: see text]-, [Formula: see text]- and methyl-[Formula: see text]-cyclodextrin in water and formamide by infrared thermal diffusion forced Rayleigh scattering (IR-TDFRS). In water the Soret coefficient, S T, of [Formula: see text]-, [Formula: see text]- and [Formula: see text]-cyclodextrin increases with increasing temperature and shows a sign change from negative to positive around T = 35 (°) C, while S T of methyl-[Formula: see text]-cyclodextrin is positive in the entire investigated temperature. In formamide S T-values of all cyclodextrins coincide and show a slight decrease with temperature. We discuss the obtained results and relate the S T-values to the different hydrogen bonding capabilities of the cyclodextrins and the used solvents. It turns out that the change of S T with temperature correlates with the partition coefficient, logP, which indicates that more hydrophilic substances show a more pronounced temperature sensitivity of S T. Additionally we obtained a surprising result measuring the refractive index contrast factor with temperature, [Formula: see text] of cyclodextrins in formamide, which might be explained by a complex formation between cyclodextrins and formamide.

  12. Historical aspects of human milk oligosaccharides.

    PubMed

    Kunz, Clemens

    2012-05-01

    This review focuses on important observations regarding infant health around 1900 when breastfeeding was not considered a matter of importance. The discovery of lactobacilli and bifidobacteria and their relevance for health and disease was an important milestone leading to a decrease in infant mortality in the first year of life. At the same time, pediatricians realized that the fecal composition of breast-fed and bottle-fed infants differed. Observations indicated that this difference is linked to milk composition, particularly due to the milk carbohydrate fraction. Circa 1930, a human milk carbohydrate fraction called gynolactose was identified. This was the starting point of research on human milk oligosaccharides (HMO). In the following years, the first HMO were identified and their functions investigated. Studies after 1950 focused on the identification of various HMO as the bifidus factor in human milk. In the following 30 years, a tremendous amount of research was done with regard to the characterization of individual HMO and HMO patterns in milk. In this short introduction to the history of HMO research, which ends circa 1980, some outstanding scientists in pediatrics and chemistry and their pioneering contributions to research in the field of HMO are presented.

  13. The functional biology of human milk oligosaccharides.

    PubMed

    Bode, Lars

    2015-11-01

    Human milk oligosaccharides (HMOs) are a group of complex sugars that are highly abundant in human milk, but currently not present in infant formula. More than a hundred different HMOs have been identified so far. The amount and composition of HMOs are highly variable between women, and each structurally defined HMO might have a distinct functionality. HMOs are not digested by the infant and serve as metabolic substrates for select microbes, contributing to shape the infant gut microbiome. HMOs act as soluble decoy receptors that block the attachment of viral, bacterial or protozoan parasite pathogens to epithelial cell surface sugars, which may help prevent infectious diseases in the gut and also the respiratory and urinary tracts. HMOs are also antimicrobials that act as bacteriostatic or bacteriocidal agents. In addition, HMOs alter host epithelial and immune cell responses with potential benefits for the neonate. The article reviews current knowledge as well as future challenges and opportunities related to the functional biology of HMOs. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

  14. Historical Aspects of Human Milk Oligosaccharides1234

    PubMed Central

    Kunz, Clemens

    2012-01-01

    This review focuses on important observations regarding infant health around 1900 when breastfeeding was not considered a matter of importance. The discovery of lactobacilli and bifidobacteria and their relevance for health and disease was an important milestone leading to a decrease in infant mortality in the first year of life. At the same time, pediatricians realized that the fecal composition of breast-fed and bottle-fed infants differed. Observations indicated that this difference is linked to milk composition, particularly due to the milk carbohydrate fraction. Circa 1930, a human milk carbohydrate fraction called gynolactose was identified. This was the starting point of research on human milk oligosaccharides (HMO). In the following years, the first HMO were identified and their functions investigated. Studies after 1950 focused on the identification of various HMO as the bifidus factor in human milk. In the following 30 years, a tremendous amount of research was done with regard to the characterization of individual HMO and HMO patterns in milk. In this short introduction to the history of HMO research, which ends circa 1980, some outstanding scientists in pediatrics and chemistry and their pioneering contributions to research in the field of HMO are presented. PMID:22585922

  15. Milk Oligosaccharides and Metabolism in Infants12

    PubMed Central

    Rudloff, Silvia; Kunz, Clemens

    2012-01-01

    Since the discovery of human milk oligosaccharides (HMO) >60 y ago, research has faced major challenges including (i) the development of methods to identify and characterize these components, (ii) the need to use HMO fractions for functional studies because single HMO were not available, (iii) the uncertainty of the purity of HMO fractions that were often “contaminated” by remainders of lactose, proteins, or glycoproteins, and (iv) the low availability of large quantities of a single HMO for animal and human studies. In the past 10 years, there has been tremendous progress in all of these areas, particularly in the development of methods for detailed structural analysis in extremely low milk volumes. The greatest success, however, is that biotechnological means are available today to produce large amounts even of a single HMO in a purity that allows human studies to be performed in the future. In this review, we summarize the current knowledge about the metabolic aspects of HMO in infants starting with the first studies by Lundblad and co-workers in the early 1980s. After discussing newer observations in recent years, the review closes with a perspective on some important questions regarding metabolic and functional aspects of HMO. PMID:22585918

  16. RP-UHPLC-UV-ESI-MS/MS analysis of LPMO generated C4-oxidized gluco-oligosaccharides after non-reductive labeling with 2-aminobenzamide.

    PubMed

    Frommhagen, Matthias; van Erven, Gijs; Sanders, Mark; van Berkel, Willem J H; Kabel, Mirjam A; Gruppen, Harry

    2017-08-07

    Lytic polysaccharide monooxygenases (LPMOs) are able to cleave recalcitrant polysaccharides, such as cellulose, by oxidizing the C1 and/or C4 atoms. The analysis of the resulting products requires a variety of analytical techniques. Up to now, these techniques mainly focused on the identification of non-oxidized and C1-oxidized oligosaccharides. The analysis of C4-oxidized gluco-oligosaccharides is mostly performed by using high pressure anion exchange chromatography (HPAEC). However, the alkaline conditions used during HPAEC analysis lead to tautomerization of C4-oxidized gluco-oligosaccharides, which limits the use of this technique. Here, we describe the use of reverse phase-ultra high performance liquid chromatography (RP-UHPLC) in combination with non-reductive 2-aminobenzamide (2-AB) labeling. Non-reductive 2-AB labeling enabled separation of C4-oxidized gluco-oligosaccharides from their non-oxidized counterparts. Moreover, RP-UHPLC does not require buffered mobile phases, which reduce mass spectrometry (MS) sensitivity. The latter is seen as an advantage over other techniques such as hydrophilic interaction liquid chromatography and porous graphitized carbon coupled to MS. RP-UHPLC coupled to UV detection and mass spectrometry allowed the identification of both labeled non-oxidized and C4-oxidized oligosaccharides. Non-reductive labeling kept the ketone at the C4-position of LPMO oxidized oligosaccharides intact, while selective reducing agents such as sodium triacetoxyborohydride (STAB) reduced this ketone group. Our results show that RP-UHPLC-UV-ESI-MS in combination with non-reductively 2-AB labeling is a suitable technique for the separation and identification of LPMO-generated C4-oxidized gluco-oligosaccharides. Copyright © 2017 The Authors. Published by Elsevier Ltd.. All rights reserved.

  17. The Ability of Bifidobacteria To Degrade Arabinoxylan Oligosaccharide Constituents and Derived Oligosaccharides Is Strain Dependent

    PubMed Central

    Rivière, Audrey; Moens, Frédéric; Selak, Marija; Maes, Dominique; Weckx, Stefan

    2014-01-01

    Arabinoxylan oligosaccharides (AXOS) are prebiotic carbohydrates with promising health-promoting properties that stimulate the activity of specific colon bacteria, in particular bifidobacteria. However, the mechanisms by which bifidobacterial strains break down these compounds in the colon is still unknown. This study investigates AXOS consumption of a large number of bifidobacterial strains (36), belonging to 11 different species, systematically. To determine their degradation mechanisms, all strains were grown on a mixture of arabinose and xylose, xylo-oligosaccharides, and complex AXOS molecules as the sole added energy sources. Based on principal component and cluster analyses of their different arabinose substituent and/or xylose backbone consumption patterns, five clusters that were species independent could be distinguished among the bifidobacterial strains tested. In parallel, the strains were screened for the presence of genes encoding several putative AXOS-degrading enzymes, but no clear-cut correlation could be made with the different degradation mechanisms. The intra- and interspecies differences in the consumption patterns of AXOS indicate that bifidobacterial strains could avoid competition among each other or even could cooperate jointly to degrade these complex prebiotics. The knowledge gained on the AXOS degradation mechanisms in bifidobacteria can be of importance in the rational design of prebiotics with tailor-made composition and thus increased specificity in the colon. PMID:24141124

  18. In vitro fermentation of oat bran obtained by debranning with a mixed culture of human fecal bacteria.

    PubMed

    Kedia, Gopal; Vázquez, José A; Charalampopoulos, Dimitris; Pandiella, Severino S

    2009-04-01

    The prebiotic potential of oat samples was investigated by in vitro shaker-flask anaerobic fermentations with human fecal cultures. The oat bran fraction was obtained by debranning and was compared with other carbon sources such as whole oat flour, glucose, and fructo-oligosaccharide. The oat bran fraction showed a decrease in culturable anaerobes and clostridia and an increase in bifidobacteria and lactobacilli populations. A similar pattern was observed in fructo-oligosaccharide. Butyrate production was higher in oat bran compared to glucose and similar to that in fructo-oligosaccharide. Production of propionate was higher in the two oat media than in fructo-oligosaccharide and glucose, which can be used as energy source by the liver. This study suggests that the oat bran fraction obtained by debranning is digested by the gut ecosystem and increases the population of beneficial bacteria in the indigenous gut microbiota. This medium also provides an energy source preferred by colonocytes when it is metabolized by the gut flora.

  19. In-Depth Characterization of N-Linked Oligosaccharides Using Fluoride-Mediated Negative Ion Microfluidic Chip LC-MS

    PubMed Central

    Ni, Wenqin; Bones, Jonathan; Karger, Barry L.

    2013-01-01

    Characterization of N-glycans by liquid chromatography-positive electrospray ionization (ESI) tandem mass spectrometry (LC-MS/MS) using a microfluidic chip packed with porous graphitized carbon (PGC) represents a rapidly developing area in oligosaccharide analysis. Positive ion ESI-MS generates B/Y-type glycosidic fragment ions under collisional induced dissociation (CID). Although these ions facilitate glycan sequencing, they provide little information on linkage and positional isomers. Isomer identification in these cases is by retention on the PGC stationary phase where the specific structural isomers can, in principle, be separated. In this paper, we broaden the applicability of the PGC microfluidic chip/MS platform by implementing fluoride-mediated negative ESI-MS. Ammonium fluoride, added to the mobile phase, aids in the formation of pseudomolecular oligosaccharide anions due to the ability of fluoride to abstract a proton from the glycan structure. The negative charge results in the generation of C-type glycosidic fragments, highly informative A-type cross ring fragment ions and additional gas phase ion reaction products (e.g., D- and E-type ions), which, when combined, lead to in-depth oligosaccharide characterization, including linkage and positional isomers. Due to the separation of anomers by the PGC phase, comparison of oligosaccharides with an intact reducing terminus to their corresponding alditols was performed, revealing a more sensitive MS and, especially, MS/MS response from the glycans with a free reducing end. Fluoride also ensured recovery of charged oligosaccharides from the PGC stationary phase. Application to the characterization of N-glycans released from polyclonal human and murine serum IgG is presented to demonstrate the effectiveness of the chip/negative ESI approach. PMID:23398125

  20. Feruloyl oligosaccharides stimulate the growth of Bifidobacterium bifidum.

    PubMed

    Yuan, Xiaoping; Wang, Jing; Yao, Huiyuan

    2005-08-01

    Insoluble dietary fiber from wheat bran contains some feruloyl groups linked to the arabinose residues in the cell wall arabinoxylan. Treatment of wheat bran insoluble dietary fiber with xylanase from Bacillus subtilis yielded feruloyl oligosacchairdes, which were purified with Amberlite XAD-2. Saponification of the feruloyl oligosaccharides released ferulic acid and arabinoxylan oligosaccharides which consist of arabinose and xylose. The effect of the feruloyl oligosacchairdes on the growth of Bifidobacterium bifidum F-35 was investigated in vitro. The B. bifidum produced acid when cultivated anaerobically in TPY broth with 0.5% feruloyl oligosacchairdes as the carbohydrate source. The biomass yield of the B. bifidum increased with increasing the concentration of feruloyl oligosaccharides in TPY broth. The maximum cell growth was increased by 50% in TPY broth supplemented with 0.1% feruloyl oligosaccharides compared to TPY broth. These results indicated that the growth of B. bifidum F-35 was promoted by the feruloyl oligosaccharides from wheat bran insoluble dietary fiber, and not suppressed by the ferulic acid moiety of them.

  1. Effect of feeding a selected combination of galacto-oligosaccharides and a strain of Bifidobacterium pseudocatenulatum on the intestinal microbiota of cats.

    PubMed

    Biagi, Giacomo; Cipollini, Irene; Bonaldo, Alessio; Grandi, Monica; Pompei, Anna; Stefanelli, Claudio; Zaghini, Giuliano

    2013-01-01

    To evaluate the growth kinetics of a strain of Bifidobacterium pseudocatenulatum (BP) on 4 oligo- or polysaccharides and the effect of feeding a selected probiotic-prebiotic combination on intestinal microbiota in cats. 10 healthy adult cats. Growth kinetics of a strain of cat-origin BP (BP-B82) on fructo-oligosaccharides, galacto-oligosaccharides (GOS), lactitol, or pectins was determined, and the combination of GOS and BP-B82 was selected. Cats received supplemental once-daily feeding of 1% GOS-BP-B82 (10(10) CFUs/d) for 15 days; fecal samples were collected for analysis the day before (day 0) and 1 and 10 days after the feeding period (day 16 and 25, respectively). Compared with the prefeeding value, mean fecal ammonia concentration was significantly lower on days 16 and 25 (288 and 281 μmol/g of fecal dry matter [fDM], respectively, vs 353 μmol/g of fDM); fecal acetic acid concentration was higher on day 16 (171 μmol/g of fDM vs 132 μmol/g of fDM). On day 16, fecal concentrations of lactic, n-valeric, and isovaleric acids (3.61, 1.52, and 3.55 μmol/g of fDM, respectively) were significantly lower than on days 0 (5.08, 18.4, and 6.48 μmol/g of fDM, respectively) and 25 (4.24, 17.3, and 6.17 μmol/g of fDM, respectively). A significant increase in fecal bifidobacteria content was observed on days 16 and 25 (7.98 and 7.52 log(10) CFUs/g of fDM, respectively), compared with the prefeeding value (5.63 log(10) CFUs/g of fDM). Results suggested that feeding 1% GOS-BP-B82 combination had some positive effects on the intestinal microbiota in cats.

  2. Utilization of major fucosylated and sialylated human milk oligosaccharides by isolated human gut microbes.

    PubMed

    Yu, Zhuo-Teng; Chen, Ceng; Newburg, David S

    2013-11-01

    Human milk oligosaccharides (HMOS) are not digested in the proximal intestine. In distal intestine, HMOS collectively modify the microbiota, but the response of individual bacteria to individual components of the HMOS is not well defined. Here, each of 25 major isolates of the human intestinal microbiota was fed individual major fucosylated and sialylated HMOS in anaerobic culture. This allowed for an assessment of the influence of specific HMOS on the growth and metabolic products of individual microbiota bacteria. Most Bifidobacteria spp. and Bacteroides spp. grew, induced α-L-fucosidase activity, and produced abundant lactate or short-chain fatty acids (SCFAs) when fed 2'-fucosyllactose (2'-FL), 3-FL, and lactodifucotetraose (LDFT). Lactobacillus delbrueckii ATCC7830, Enterococcus faecalis ATCC19433, and Streptococcus thermophilus ATCC19258 exhibited slight growth, pH reduction, and lactate production when supplemented with 2'-FL or 3-FL, but not LDFT. Supplementation with 3'-sialyllactose (3'-SL) and 6'-SL promoted moderate growth of Bifidobacterium longum JCM7007, 7009, 7010, 7011, 1272, 11347, ATCC15708, Bacteroides vulgatus ATCC8482, and B. thetaiotaomicron ATCC29148; accordingly, these bacteria exhibited greater neuraminidase activity and produced copious lactate, SCFA, or both. Lactobacillus delbrueckii ATCC7830 also consumed 6'-SL. In contrast, Clostridium spp., L. rhamnosus ATCC53103, E. faecalis ATCC29200, Staphylococcus spp., Enterobacter spp., and Escherichia coli K12 did not consume milk oligosaccharides nor produce appreciable acidic fermentation products. Specific Bifidobacteria and Bacteroides differentially digest specific individual HMOS, with the major fucosylated milk oligosaccharides most strongly stimulating key species of mutualist symbionts. This suggests strategies for treating dysbiosis of the microbiota and associated inflammatory disorders.

  3. Utilization of major fucosylated and sialylated human milk oligosaccharides by isolated human gut microbes

    PubMed Central

    Yu, Zhuo-Teng; Chen, Ceng; Newburg, David S

    2013-01-01

    Human milk oligosaccharides (HMOS) are not digested in the proximal intestine. In distal intestine, HMOS collectively modify the microbiota, but the response of individual bacteria to individual components of the HMOS is not well defined. Here, each of 25 major isolates of the human intestinal microbiota was fed individual major fucosylated and sialylated HMOS in anaerobic culture. This allowed for an assessment of the influence of specific HMOS on the growth and metabolic products of individual microbiota bacteria. Most Bifidobacteria spp. and Bacteroides spp. grew, induced α-l-fucosidase activity, and produced abundant lactate or short-chain fatty acids (SCFAs) when fed 2′-fucosyllactose (2′-FL), 3-FL, and lactodifucotetraose (LDFT). Lactobacillus delbrueckii ATCC7830, Enterococcus faecalis ATCC19433, and Streptococcus thermophilus ATCC19258 exhibited slight growth, pH reduction, and lactate production when supplemented with 2′-FL or 3-FL, but not LDFT. Supplementation with 3′-sialyllactose (3′-SL) and 6′-SL promoted moderate growth of Bifidobacterium longum JCM7007, 7009, 7010, 7011, 1272, 11347, ATCC15708, Bacteroides vulgatus ATCC8482, and B. thetaiotaomicron ATCC29148; accordingly, these bacteria exhibited greater neuraminidase activity and produced copious lactate, SCFA, or both. Lactobacillus delbrueckii ATCC7830 also consumed 6′-SL. In contrast, Clostridium spp., L. rhamnosus ATCC53103, E. faecalis ATCC29200, Staphylococcus spp., Enterobacter spp., and Escherichia coli K12 did not consume milk oligosaccharides nor produce appreciable acidic fermentation products. Specific Bifidobacteria and Bacteroides differentially digest specific individual HMOS, with the major fucosylated milk oligosaccharides most strongly stimulating key species of mutualist symbionts. This suggests strategies for treating dysbiosis of the microbiota and associated inflammatory disorders. PMID:24013960

  4. A C4-oxidizing Lytic Polysaccharide Monooxygenase Cleaving Both Cellulose and Cello-oligosaccharides*

    PubMed Central

    Isaksen, Trine; Westereng, Bjørge; Aachmann, Finn L.; Agger, Jane W.; Kracher, Daniel; Kittl, Roman; Ludwig, Roland; Haltrich, Dietmar; Eijsink, Vincent G. H.; Horn, Svein J.

    2014-01-01

    Lignocellulosic biomass is a renewable resource that significantly can substitute fossil resources for the production of fuels, chemicals, and materials. Efficient saccharification of this biomass to fermentable sugars will be a key technology in future biorefineries. Traditionally, saccharification was thought to be accomplished by mixtures of hydrolytic enzymes. However, recently it has been shown that lytic polysaccharide monooxygenases (LPMOs) contribute to this process by catalyzing oxidative cleavage of insoluble polysaccharides utilizing a mechanism involving molecular oxygen and an electron donor. These enzymes thus represent novel tools for the saccharification of plant biomass. Most characterized LPMOs, including all reported bacterial LPMOs, form aldonic acids, i.e., products oxidized in the C1 position of the terminal sugar. Oxidation at other positions has been observed, and there has been some debate concerning the nature of this position (C4 or C6). In this study, we have characterized an LPMO from Neurospora crassa (NcLPMO9C; also known as NCU02916 and NcGH61–3). Remarkably, and in contrast to all previously characterized LPMOs, which are active only on polysaccharides, NcLPMO9C is able to cleave soluble cello-oligosaccharides as short as a tetramer, a property that allowed detailed product analysis. Using mass spectrometry and NMR, we show that the cello-oligosaccharide products released by this enzyme contain a C4 gemdiol/keto group at the nonreducing end. PMID:24324265

  5. Novel arabinan and galactan oligosaccharides from dicotyledonous plants

    NASA Astrophysics Data System (ADS)

    Wefers, Daniel; Tyl, Catrin; Bunzel, Mirko

    2014-11-01

    Arabinans and galactans are neutral pectic side chains and an important part of the cell walls of dicotyledonous plants. To get a detailed insight into their fine structure, various oligosaccharides were isolated from quinoa, potato galactan, and sugar beet pulp after enzymatic treatment. LC-MS2 and one- and two-dimensional NMR spectroscopy were used for unambiguous structural characterization. It was demonstrated that arabinans contain β-(1→3)-linked arabinobiose as a side chain in quinoa seeds, while potato galactan was comprised of β-(1→4)-linked galactopyranoses which are interspersed with α-(1→4)-linked arabinopyranoses. Additionally, an oligosaccharide with two adjacent arabinofuranose units O2-substituted with two ferulic acid monomers was characterized. The isolated oligosaccharides gave further insight into the structures of pectic side chains and may have an impact on plant physiology and dietary fiber fermentation.

  6. Natural variability in bovine milk oligosaccharides from Danish Jersey and Holstein-Friesian breeds

    PubMed Central

    Sundekilde, Ulrik K; Barile, Daniela; Meyrand, Mickael; Poulsen, Nina A; Larsen, Lotte B; Lebrilla, Carlito B.; Bruce, German J.; Bertram, Hanne C

    2012-01-01

    Free oligosaccharides are key components of human milk and play multiple roles in the health of the neonate, by stimulating growth of selected beneficial bacteria in the gut, participating in development of the brain and exerting anti-pathogenic activity. However, the concentration of oligosaccharides is low in mature bovine milk, normally used for infant formula, compared with both human colostrum and mature human milk. Characterization of bovine milk oligosaccharides in different breeds is crucial for the identification of viable sources for oligosaccharide purification. An improved source of oligosaccharides can lead to infant formula with improved oligosaccharide functionality. In the present study we have analyzed milk oligosaccharides by high-performance liquid chromatography chip quadrupole time-of-flight mass spectrometry and performed a detailed data analysis using both univariate and multivariate methods. Both statistical tools revealed several differences in oligosaccharide profiles between milk samples from the two Danish breeds; Jersey and Holstein-Friesians. Jersey milk contained higher relative amounts of both sialylated and the more complex neutral fucosylated oligosaccharides, while the Holstein-Friesian milk had higher abundance of smaller and simpler neutral oligosaccharides. The statistical analyses revealed that Jersey milk contain significantly higher levels of fucosylated oligosaccharides than Holstein-Friesian milk. Jersey milk also possesses oligosaccharides with a higher degree of complexity and functional residues (fucose and sialic acid) suggesting it may therefore offer advantages in term of a wider array of bioactivities. PMID:22632419

  7. Novel strategy for herbal species classification based on UPLC-HRMS oligosaccharide profiling.

    PubMed

    Tie, Cai; Hu, Ting; Guo, Baolin; Zhang, Jinlan

    2015-01-01

    Oligosaccharides, which exist widely in herbs, present diverse important pharmacological activities. However, the complexity of oligosaccharides seriously challenges their profiling, quality control, and elucidation of activity. In this paper, a novel oligosaccharide analytical method based on a new derivatization pretreatment and ultra-performance liquid chromatography coupled with high resolution tandem mass spectrometry (UPLC-HRMS) procedure was developed to rapidly profile and identify the oligosaccharides of Epimedium. Oligosaccharides are easily derivatized by 2,4-bis(diethylamino)-6-hydrazino-1,3,5-triazine under convenient and mild conditions. Without any further purification steps, oligosaccharides were analyzed by an established UPLC-HRMS/MS method with high sensitivity, good separation efficiency and speed. Benefitting from the derivatization, the oligosaccharides generated a response in the MS two orders of magnitude higher than that of the free oligosaccharide. Oligosaccharides of 52 Epimedium samples were profiled and identified based on the high-resolution mass spectral data. A total of 66 oligosaccharide compounds detected in 52 Epimedium herbs were relatively quantified and statistically processed by principal component analysis (PCA). The 52 Epimedium herbs could be classified into different species based on their oligosaccharide composition and content. Thirteen oligosaccharide compounds demonstrated potential as markers for Epimedium species classification, and their structures were preliminarily identified using MS/MS spectra.

  8. Synthesis of sulfated oligosaccharides by cystic fibrosis trachea epithelial cells.

    PubMed

    Mendicino, J; Sangadala, S

    1999-11-01

    The mucin glycoproteins in tracheal mucus of patients with cystic fibrosis is more highly sulfated than the corresponding secretions from healthy individuals [16]. In order to further characterize these differences in sulfation and possibly also glycosylation patterns, we compared the structures of sulfated mucin oligosaccharides synthesized by continuously cultured human tracheal cells transformed by simian virus 40. The synthesis of highly sulfated oligosaccharide chains in mucins secreted by normal human epithelial and submucosal cell lines were compared with mucins formed by cystic fibrosis tracheal epithelial and submucosal cell lines. The epithelial cell lines from cystic fibrosis trachea showed a higher rate of sulfate uptake and a significantly higher rate of synthesis and sulfation of high molecular weight chains. Mucins synthesized by each cell line in the presence of 35SO4 were isolated and oligosaccharide chains were released by beta-elimination and separated by ion exchange chromatography and gel filtration. The sulfated high molecular weight chains synthesized by the cystic fibrosis cell lines were characterized by methylation analysis and sequential glycosidase digestion before and after desulfation. Carbohydrate analysis yielded Fuc, Gal and GlcNAc in a ratio of 1:2:2.2 and only one galactosaminitol residue for about every 150-200 sugar residues present. The average molecular size of oligosaccharide chains in these fractions was between 30,000-40,000 daltons. These studies show that increased sulfation of oligosaccharides in mucins synthesized by cells from cystic fibrosis trachea is accompanied by a significant increase in the extension of a basic branched structure present in many of the lower molecular weight oligosaccharides.

  9. Purification and characterization of a xyloglucan oligosaccharide-specific xylosidase from pea seedlings

    SciTech Connect

    O'Neill, R.A.; Albersheim, P.; Darvill, A.G. )

    1989-12-05

    An {alpha}-xylosidase that acts on oligosaccharide fragments of xyloglucan, a plant cell wall polysaccharide, was purified from pea (Pisum sativum) epicotyls that had been treated with an auxin analog. The enzyme had an apparent molecular mass of 85,000 Da according to sodium dodecyl sulfate-polyacrylamide gel electrophoresis and 79,000 Da according to gel-permeation chromatography under nondenaturing conditions. The purified xylosidase consisted of a series of closely related, enzymatically active proteins with isoelectric points ranging from about pH 7.35 to 7.7; the xylosidases were separated by chromatofocusing. The pH optimum of the mixed xylosidase was 4.9-5.1. The substrate specificity of the xylosidase mixture was determined by purification and structural characterization of the products of treating xyloglucan-oligosaccharide substrates with the enzyme. Characterization of the substrates and products included elution volume from a gel-permeation column, glycosyl residue and glycosyl linkage composition analyses, fast atom bombardment-mass spectrometry, and {sup 1}H NMR spectroscopy. The enzyme specifically cleaved only one of the {alpha}-xylosidic linkages of xyloglucan-oligosaccharide substrates, the one attached to a 6-linked glucosyl residue, not those attached to the 4,6-linked glucosyl residues. The enzyme was unable to cleave the xylosidic linkage of p-nitrophenyl-{alpha}-D-xylopyranoside or the {alpha}-xylosidic linkage to C-6 of glucose in the disaccharide isoprimeverose. The enzyme was also unable to release measurable amounts of xylose from large xyloglucan polymers.

  10. A new, quick, highly sensitive ultramicro-analysis method for the identification of fructose removed from fructofuranosyl-containing gluco-oligosaccharides by ESI-CID-MS/MS.

    PubMed

    Zhang, Hong-Tao; Zhu, Li; Zhang, Shuang; Zhan, Xiao-Bei; Lin, Chi-Chung

    2014-10-29

    An efficient, highly sensitive, and ultramicroscale analytical method for the identification of fructose removed from fructofuranosyl-containing gluco-oligosaccharides, including malto-oligosyl fructofuranosides and oligomeric (1→2)-α-D-glucopyranosyl-(1→2)-β-D-fructofuranosides by ESI-CID-MS/MS has been developed with proven applications far superior to the existing method using NMR. With the established principle of diagnostic fragmentation by ESI-CID-MS/MS, the terminal saccharide (either glucose or fructose) can be readily and unambiguously determined at high sensitivity without a tedious derivatization process. Detection of the A-type fragmentation (0,4)A-h type ion, and (0,2)A type ion are useful as a diagnostic fragmentation tool to identify whether fructose terminal is removed from oligosaccharides. It will facilitate the efficient production of suitable oligosaccharide microarrays crucial for studies on carbohydrate-protein interaction in seeking functional carbohydrates.

  11. Evolved β-Galactosidases from Geobacillus stearothermophilus with Improved Transgalactosylation Yield for Galacto-Oligosaccharide Production▿ †

    PubMed Central

    Placier, Gaël; Watzlawick, Hildegard; Rabiller, Claude; Mattes, Ralf

    2009-01-01

    A mutagenesis approach was applied to the β-galactosidase BgaB from Geobacillus stearothermophilus KVE39 in order to improve its enzymatic transglycosylation of lactose into oligosaccharides. A simple screening strategy, which was based on the reduction of the hydrolysis of a potential transglycosylation product (lactosucrose), provided mutant enzymes possessing improved synthetic properties for the autocondensation product from nitrophenyl-galactoside and galacto-oligosaccharides (GOS) from lactose. The effects of the mutations on enzyme activity and kinetics were determined. An change of one arginine to lysine (R109K) increased the oligosaccharide yield compared to that for the wild-type BgaB. Subsequently, saturation mutagenesis at this position demonstrated that valine and tryptophan further increased the transglycosylation performance of BgaB. During the transglycosylation reaction with lactose of the evolved β-galactosidases, a major trisaccharide was formed. Its structure was characterized as β-d-galactopyranosyl-(1→3)-β-d-galactopyranosyl-(1→4)-d-glucopyranoside (3′-galactosyl-lactose). At the lactose concentration of 18% (wt/vol), this trisaccharide was obtained in yields of 11.5% (wt/wt) with GP21 (BgaB R109K), 21% with GP637.2 (BgaB R109V), and only 2% with the wild-type BgaB enzyme. GP643.3 (BgaB R109W) was shown to be the most efficient mutant, with a 3′-galactosyl-lactose production of 23%. PMID:19666723

  12. Transcriptome Profiling of Bovine Milk Oligosaccharide Metabolism Genes Using RNA-Sequencing

    PubMed Central

    Wickramasinghe, Saumya; Hua, Serenus; Rincon, Gonzalo; Islas-Trejo, Alma; German, J. Bruce; Lebrilla, Carlito B.; Medrano, Juan F.

    2011-01-01

    This study examines the genes coding for enzymes involved in bovine milk oligosaccharide metabolism by comparing the oligosaccharide profiles with the expressions of glycosylation-related genes. Fresh milk samples (n = 32) were collected from four Holstein and Jersey cows at days 1, 15, 90 and 250 of lactation and free milk oligosaccharide profiles were analyzed. RNA was extracted from milk somatic cells at days 15 and 250 of lactation (n = 12) and gene expression analysis was conducted by RNA-Sequencing. A list was created of 121 glycosylation-related genes involved in oligosaccharide metabolism pathways in bovine by analyzing the oligosaccharide profiles and performing an extensive literature search. No significant differences were observed in either oligosaccharide profiles or expressions of glycosylation-related genes between Holstein and Jersey cows. The highest concentrations of free oligosaccharides were observed in the colostrum samples and a sharp decrease was observed in the concentration of free oligosaccharides on day 15, followed by progressive decrease on days 90 and 250. Ninety-two glycosylation-related genes were expressed in milk somatic cells. Most of these genes exhibited higher expression in day 250 samples indicating increases in net glycosylation-related metabolism in spite of decreases in free milk oligosaccharides in late lactation milk. Even though fucosylated free oligosaccharides were not identified, gene expression indicated the likely presence of fucosylated oligosaccharides in bovine milk. Fucosidase genes were expressed in milk and a possible explanation for not detecting fucosylated free oligosaccharides is the degradation of large fucosylated free oligosaccharides by the fucosidases. Detailed characterization of enzymes encoded by the 92 glycosylation-related genes identified in this study will provide the basic knowledge for metabolic network analysis of oligosaccharides in mammalian milk. These candidate genes will guide

  13. Role of human milk oligosaccharides in Group B Streptococcus colonisation.

    PubMed

    Andreas, Nicholas J; Al-Khalidi, Asmaa; Jaiteh, Mustapha; Clarke, Edward; Hyde, Matthew J; Modi, Neena; Holmes, Elaine; Kampmann, Beate; Mehring Le Doare, Kirsty

    2016-08-01

    Group B Streptococcus (GBS) infection is a major cause of morbidity and mortality in infants. The major risk factor for GBS disease is maternal and subsequent infant colonisation. It is unknown whether human milk oligosaccharides (HMOs) protect against GBS colonisation. HMO production is genetically determined and linked to the Lewis antigen system. We aimed to investigate the association between HMOs and infant GBS colonisation between birth and postnatal day 90. Rectovaginal swabs were collected at delivery, as well as colostrum/breast milk, infant nasopharyngeal and rectal swabs at birth, 6 days and days 60-89 postpartum from 183 Gambian mother/infant pairs. GBS colonisation and serotypes were determined using culture and PCR. (1)H nuclear magnetic resonance spectroscopy was used to characterise the mother's Lewis status and HMO profile in breast milk. Mothers who were Lewis-positive were significantly less likely to be colonised by GBS (X (2)=12.50, P<0.001). Infants of Lewis-positive mothers were less likely GBS colonised at birth (X (2)=4.88 P=0.03) and more likely to clear colonisation between birth and days 60-89 than infants born to Lewis-negative women (P=0.05). There was no association between Secretor status and GBS colonisation. In vitro work revealed that lacto-N-difucohexaose I (LNDFHI) correlated with a reduction in the growth of GBS. Our results suggest that HMO such as LNDFHI may be a useful adjunct in reducing maternal and infant colonisation and hence invasive GBS disease. Secretor status offers utility as a stratification variable in GBS clinical trials.

  14. Role of human milk oligosaccharides in Group B Streptococcus colonisation

    PubMed Central

    Andreas, Nicholas J; Al-Khalidi, Asmaa; Jaiteh, Mustapha; Clarke, Edward; Hyde, Matthew J; Modi, Neena; Holmes, Elaine; Kampmann, Beate; Mehring Le Doare, Kirsty

    2016-01-01

    Group B Streptococcus (GBS) infection is a major cause of morbidity and mortality in infants. The major risk factor for GBS disease is maternal and subsequent infant colonisation. It is unknown whether human milk oligosaccharides (HMOs) protect against GBS colonisation. HMO production is genetically determined and linked to the Lewis antigen system. We aimed to investigate the association between HMOs and infant GBS colonisation between birth and postnatal day 90. Rectovaginal swabs were collected at delivery, as well as colostrum/breast milk, infant nasopharyngeal and rectal swabs at birth, 6 days and days 60–89 postpartum from 183 Gambian mother/infant pairs. GBS colonisation and serotypes were determined using culture and PCR. 1H nuclear magnetic resonance spectroscopy was used to characterise the mother's Lewis status and HMO profile in breast milk. Mothers who were Lewis-positive were significantly less likely to be colonised by GBS (X2=12.50, P<0.001). Infants of Lewis-positive mothers were less likely GBS colonised at birth (X2=4.88 P=0.03) and more likely to clear colonisation between birth and days 60–89 than infants born to Lewis-negative women (P=0.05). There was no association between Secretor status and GBS colonisation. In vitro work revealed that lacto-N-difucohexaose I (LNDFHI) correlated with a reduction in the growth of GBS. Our results suggest that HMO such as LNDFHI may be a useful adjunct in reducing maternal and infant colonisation and hence invasive GBS disease. Secretor status offers utility as a stratification variable in GBS clinical trials. PMID:27588204

  15. Molecular diversity of the genetic loci responsible for lipopolysaccharide core oligosaccharide assembly within the genus Salmonella.

    PubMed

    Kaniuk, Natalia A; Monteiro, Mario A; Parker, Craig T; Whitfield, Chris

    2002-12-01

    The waa locus on the chromosome of Salmonella enterica encodes enzymes involved in the assembly of the core oligosaccharide region of the lipopolysaccharide (LPS) molecule. To date, there are two known core structures in Salmonella, represented by serovars Typhimurium (subspecies I) and Arizonae (subspecies IIIA). The waa locus for serovar Typhimurium has been characterized. Here, the corresponding locus from serovar Arizonae is described, and the molecular basis for the distinctive structures is established. Eleven of the 13 open reading frames (ORFs) are shared by the two loci and encode conserved proteins of known function. Two polymorphic regions distinguish the waa loci. One involves the waaK gene, the product of which adds a terminal alpha-1,2-linked N-acetylglucosamine residue that characterizes the serovar Typhimurium core oligosaccharide. There is an extensive internal deletion within waaK of serovar Arizonae. The serovar Arizonae locus contains a novel ORF (waaH) between the waaB and waaP genes. Structural analyses and in vitro glycosyltransferase assays identified WaaH as the UDP-glucose:(glucosyl) LPS alpha-1,2-glucosyltransferase responsible for the addition of the characteristic terminal glucose residue found in serovar Arizonae. Isolates comprising the Salmonella Reference Collections, SARC (representing the eight subspecies of S. enterica) and SARB (representing subspecies I), were examined to assess the distribution of the waa locus polymorphic regions in natural populations. These comparative studies identified additional waa locus polymorphisms, shedding light on the genetic basis for diversity in the LPS core oligosaccharides of Salmonella isolates and identifying potential sources of further novel LPS structures.

  16. In Vitro Fermentation of caprine milk oligosaccharides by bifidobacteria isolated from breast-fed infants

    PubMed Central

    Thum, Caroline; Roy, Nicole C; McNabb, Warren C; Otter, Don E; Cookson, Adrian L

    2015-01-01

    This study was conducted to investigate the catabolism and fermentation of caprine milk oligosaccharides (CMO) by selected bifidobacteria isolated from 4 breast-fed infants. Seventeen bifidobacterial isolates consisting of 3 different species (Bifidobacterium breve, Bifidobacterium longum subsp. longum and Bifidobacterium bifidum) were investigated. A CMO-enriched fraction (CMOF) (50% oligosaccharides, 10% galacto-oligosaccharides (GOS), 20% lactose, 10% glucose and 10% galactose) from caprine cheese whey was added to a growth medium as a sole source of fermentable carbohydrate. The inclusion of the CMOF was associated with increased bifidobacterial growth for all strains compared to glucose, lactose, GOS, inulin, oligofructose, 3'-sialyl-lactose and 6'-sialyl-lactose. Only one B. bifidum strain (AGR2166) was able to utilize the sialyl-CMO, 3'-sialyl-lactose and 6'-sialyl-lactose, as carbohydrate sources. The inclusion of CMOF increased the production of acetic and lactic acid (P < 0.001) after 36 h of anaerobic fermentation at 37°C, when compared to other fermentable substrates. Two B. bifidum strains (AGR2166 and AGR2168) utilised CMO, contained in the CMOF, to a greater extent than B. breve or B. longum subsp longum isolates, and this increased CMO utilization was associated with enhanced sialidase activity. CMOF stimulated bifidobacterial growth when compared to other tested fermentable carbohydrates and also increased the consumption of mono- and disaccharides, such as galactose and lactose present in the CMOF. These findings indicate that the dietary consumption of CMO may stimulate the growth and metabolism of intestinal Bifidobacteria spp. including B. bifidum typically found in the large intestine of breast-fed infants. PMID:26587678

  17. Screening Substrate Properties of Microorganisms for Biosensor Detection of Oligosaccharides

    USDA-ARS?s Scientific Manuscript database

    Oligosaccharides feature high biological activity ensuring their wide application in the biotechnology, food, and cosmetic industries. On the other hand they are considered environmental pollutants. The study outlines a biosensor approach to detect these substances which is important from above st...

  18. Transcriptional analysis of oligosaccharide utilization by Bifidobacterium lactis Bl-04

    PubMed Central

    2013-01-01

    Background Probiotic bifidobacteria in combination with prebiotic carbohydrates have documented positive effects on human health regarding gastrointestinal disorders and improved immunity, however the selective routes of uptake remain unknown for most candidate prebiotics. The differential transcriptomes of Bifidobacterium animalis subsp. lactis Bl-04, induced by 11 potential prebiotic oligosaccharides were analyzed to identify the genetic loci involved in the uptake and catabolism of α- and β-linked hexoses, and β-xylosides. Results The overall transcriptome was modulated dependent on the type of glycoside (galactosides, glucosides or xylosides) utilized. Carbohydrate transporters of the major facilitator superfamily (induced by gentiobiose and β-galacto-oligosaccharides (GOS)) and ATP-binding cassette (ABC) transporters (upregulated by cellobiose, GOS, isomaltose, maltotriose, melibiose, panose, raffinose, stachyose, xylobiose and β-xylo-oligosaccharides) were differentially upregulated, together with glycoside hydrolases from families 1, 2, 13, 36, 42, 43 and 77. Sequence analysis of the identified solute-binding proteins that determine the specificity of ABC transporters revealed similarities in the breadth and selectivity of prebiotic utilization by bifidobacteria. Conclusion This study identified the differential gene expression for utilization of potential prebiotics highlighting the extensive capabilities of Bifidobacterium lactis Bl-04 to utilize oligosaccharides. Results provide insights into the ability of this probiotic microbe to utilize indigestible carbohydrates in the human gastrointestinal tract. PMID:23663691

  19. Transcriptional analysis of oligosaccharide utilization by Bifidobacterium lactis Bl-04.

    PubMed

    Andersen, Joakim M; Barrangou, Rodolphe; Abou Hachem, Maher; Lahtinen, Sampo J; Goh, Yong Jun; Svensson, Birte; Klaenhammer, Todd R

    2013-05-10

    Probiotic bifidobacteria in combination with prebiotic carbohydrates have documented positive effects on human health regarding gastrointestinal disorders and improved immunity, however the selective routes of uptake remain unknown for most candidate prebiotics. The differential transcriptomes of Bifidobacterium animalis subsp. lactis Bl-04, induced by 11 potential prebiotic oligosaccharides were analyzed to identify the genetic loci involved in the uptake and catabolism of α- and β-linked hexoses, and β-xylosides. The overall transcriptome was modulated dependent on the type of glycoside (galactosides, glucosides or xylosides) utilized. Carbohydrate transporters of the major facilitator superfamily (induced by gentiobiose and β-galacto-oligosaccharides (GOS)) and ATP-binding cassette (ABC) transporters (upregulated by cellobiose, GOS, isomaltose, maltotriose, melibiose, panose, raffinose, stachyose, xylobiose and β-xylo-oligosaccharides) were differentially upregulated, together with glycoside hydrolases from families 1, 2, 13, 36, 42, 43 and 77. Sequence analysis of the identified solute-binding proteins that determine the specificity of ABC transporters revealed similarities in the breadth and selectivity of prebiotic utilization by bifidobacteria. This study identified the differential gene expression for utilization of potential prebiotics highlighting the extensive capabilities of Bifidobacterium lactis Bl-04 to utilize oligosaccharides. Results provide insights into the ability of this probiotic microbe to utilize indigestible carbohydrates in the human gastrointestinal tract.

  20. Chitin oligosaccharide deacetylase from Shewanella baltica ATCC BAA-1091.

    PubMed

    Hirano, Takako; Shiraishi, Haruka; Ikejima, Masafumi; Uehara, Rie; Hakamata, Wataru; Nishio, Toshiyuki

    2017-03-01

    Chitin oligosaccharide deacetylase (COD) from bacteria that have been examined so far typically comprise two carbohydrate-binding domains (CBDs) and one polysaccharide deacetylase domain. In contrast, Shewanella baltica ATCC BAA-1091 COD (Sb-COD) has only one CBD, yet exhibits chitin-binding properties and substrate specificities similar to those of other CODs.

  1. New Glucuronic Acid Donors for the Modular Synthesis of Heparan Sulfate Oligosaccharides**

    PubMed Central

    Dhamale, Omkar P.; Zong, Chengli; Al-Mafraji, Kanar

    2014-01-01

    Although hundreds of heparan sulfate (HS) binding proteins have been implicated in a myriad of physiological and pathological processes, very little information is known about ligand requirements for binding and mediating biological activities by these proteins. We report here a streamlined approach for the preparation of modular disaccharide building blocks that will facilitate the assembly of libraries of HS oligosaccharides for structure-activity relationship studies. In particular, we have found that glucuronic acid donors, which usually perform poorly in glycosylations, can give high yields of coupling product when the C-2 hydroxyl is protected with a permanent 4-acetoxy-2,2-dimethyl butanoyl- (PivOAc) or temporary levulinoyl (Lev) ester and the C-4 hydroxyl modified with a selectively removable 2-methylnaphthyl (Nap) ether. It has been shown that the PivOAc ester can be removed without affecting sulfate esters making it an ideal protecting group for HS oligosaccharide assembly. Iduronic acid donors exhibit more favorable glycosyl donating properties and a compound protected with a Lev ester at C-2 and an Fmoc function at the C-4 hydroxyl gave coupling products in high yield. The new donors avoid post-glycosylation oxidation and therefore allow the facile preparation of modular disaccharide building blocks. PMID:24549353

  2. Structural differences of prebiotic oligosaccharides influence their capability to enhance iron absorption in deficient rats.

    PubMed

    Laparra, José Moisés; Díez-Municio, Marina; Herrero, Miguel; Moreno, F Javier

    2014-10-01

    This study evaluates the influence of novel galacto-oligosaccharides derived from lactulose (GOS-Lu), kojibiose or 4'-galactosyl-kojibiose in hematological parameters of Fe homeostasis using Fe-deficient animals. Liver TfR-2, IL-6, NFκB and PPAR-γ expression (mRNA) were also determined by RT-qPCR analyses, and active hepcidin peptide production and short chain fatty acids by LC coupled to MS/MS or UV detection. Feeding animals with GOS-Lu or kojibiose together with FeCl3 increased hemoglobin (Hb) production (by 17%) and mean Hb concentration into erythrocytes relative to animals administered with FeCl3 alone (14.1% and 19.7%, respectively). Animals administered with prebiotics showed decreased plasmatic hepcidin levels, contributing to a higher intestinal absorption of the micronutrient. These data indicate that concurrent administration of these potentially prebiotic oligosaccharides together with a supplement of Fe ameliorates inflammation-mediated perturbations in the liver, according to the particular structure of the prebiotic compound, and result an attractive strategy to improve Fe absorption.

  3. Thermopressurized diluted phosphoric acid pretreatment of ligno(hemi)cellulose to make free sugars and nutraceutical oligosaccharides.

    PubMed

    Tiboni, Marcela; Grzybowski, Adelia; Baldo, Gizele Rejane; Dias, Edson Flausino; Tanner, Robert D; Kornfield, Julia Ann; Fontana, José Domingos

    2014-06-01

    Ligno(hemi)cellulosics (L(h)Cs) as sugarcane bagasse and loblolly pine sawdust are currently being used to produce biofuels such as bioethanol and biobutanol through fermentation of free sugars that are often obtained enzymatically. However, this bioconversion requires a pretreatment to solubilize the hemicellulose fractions, thus facilitating the action of the cellulolytic enzymes. Instead of the main free monosaccharides used in these current models, the modulation of thermopressurized orthophosphoric acid as a pretreatment, in the ranges of 3-12 atm and pH 1.5-2.5, can produce nondigestible oligosaccharides (NDOS) such as xylo-oligosaccharides (XOS) because heteroxylan is present in both types of hardwood and softwood hemicelluloses. A comparative thin-layer chromatographic analysis of the hydrolytic products showed the best conditions for NDOS production to be 7 atm/water, pH 2.25 and 2.50, and 8.5 atm/water for both sources. Particular hydrolysates from 7 atm (171 °C) at pHs 2.25 and 2.50 both for cane bagasse and pine sawdust, with respective oligosaccharide contents of 57 and 59 %, once mixed in a proportion of 1:1 for each plant source, were used in vitro as carbon sources for Bifidobacterium or Lactobacillus. Once both bacteria attained the stationary phase of growth, an unforeseen feature emerged: the preference of B. animalis for bagasse hydrolysates and, conversely, the preference of L. casei for pine hydrolysates. Considering the fact that nutraceutical oligosaccharides from both hemicelluloses correspond to higher value-added byproducts, the technology using a much diluted thermopressurized orthophosphoric acid pretreatment becomes an attractive choice for L(h)Cs.

  4. Chemical characterization of the oligosaccharides in Bactrian camel (Camelus bactrianus) milk and colostrum.

    PubMed

    Fukuda, K; Yamamoto, A; Ganzorig, K; Khuukhenbaatar, J; Senda, A; Saito, T; Urashima, T

    2010-12-01

    Bactrian camel milk and colostrum are commonly used as foods in Mongolia, whose people believe that these products promote human health. It has been hypothesized that milk oligosaccharides are biologically significant components of human milk, acting as receptor analogs that inhibit the attachment of pathogenic microorganisms to the colonic mucosa, and as prebiotics, which stimulate the growth of bifidobacteria within the infant colon. To evaluate their biological significance, we studied the oligosaccharides present in samples of Bactrian camel milk and colostrum. Using (1)H-nuclear magnetic resonance spectroscopy, we identified and characterized the following oligosaccharides of camel colostrum: Gal(β1-4)[Fuc(α1-3)]Glc (3-fucosyllactose), Gal(β1-3)Gal(β1-4)Glc (3'-galactosyllactose), Gal(β1-6)Gal(β1-4)Glc (6'-galactosyllactose), Neu5Ac(α2-3)Gal(β1-4)Glc (3'-sialyllactose), Neu5Ac(α2-6)Gal(β1-4)Glc (6'-sialyllactose), Neu5Ac(α2-3)Gal(β1-3)Gal(β1-4)Glc (sialyl-3'-galactosyllactose), Neu5Ac(α2-6)Gal(β1-4)GlcNAc(β1-3)Gal(β1-4)Glc (sialyllacto-N-tetraose c), Neu5Ac(α2-3)Gal(β1-3)[Gal(β1-4)GlcNAc(β1-6)]Gal(β1-4)Glc (sialyllacto-N-novopentaose a), Gal(β1-3)[Neu5Ac(α2-6)Gal(β1-4)GlcNAc(β1-6)]Gal(β1-4)Glc (sialyllacto-N-novopentaose b); and Neu5Ac(α2-6)Gal(β1-4)GlcNAc(β1-3)[Gal(β1-4)GlcNAc(β1-6)]Gal(β1-4)Glc (monosialyllacto-N-neohexaose). The oligosaccharides in the mature camel milk were characterized as 3'-galactosyllactose, Gal(β1-3)[Gal(β1-4)GlcNAc(β1-6)]Gal(β1-4)Glc (lacto-N-novopentaose I), and 3'-sialyllactose.

  5. Lactic acid bacteria fermentation of human milk oligosaccharide components, human milk oligosaccharides and galactooligosaccharides.

    PubMed

    Schwab, Clarissa; Gänzle, Michael

    2011-02-01

    Human milk contains about 7% lactose and 1% human milk oligosaccharides (HMOs) consisting of lactose with linked fucose, N-acetylglucosamine and sialic acid. In infant formula, galactooligosaccharides (GOSs) are added to replace HMOs. This study investigated the ability of six strains of lactic acid bacteria (LAB), Lactobacillus acidophilus, Lactobacillus plantarum, Lactobacillus fermentum, Lactobacillus reuteri, Streptococcus thermophilus and Leuconostoc mesenteroides subsp. cremoris, to digest HMO components, defined HMOs, and GOSs. All strains grew on lactose and glucose. N-acetylglucosamine utilization varied between strains and was maximal in L. plantarum; fucose utilization was low or absent in all strains. Both hetero- and homofermentative LAB utilized N-acetylglucosamine via the Embden-Meyerhof pathway. Lactobacillus acidophilus and L. plantarum were the most versatile in hydrolysing pNP analogues and the only strains releasing mono- and disaccharides from defined HMOs. Whole cells of all six LAB hydrolysed oNP-galactoside and pNP-galactoside indicating β-galactosidase activity. High β-galactosidase activity of L. reuteri, L. fermentum, S. thermophilus and L. mesenteroides subsp. cremoris whole cells correlated to lactose and GOS hydrolysis. Hydrolysis of lactose and GOSs by heterologously expressed β-galactosidases confirmed that LAB β-galactosidases are involved in GOS digestion. In summary, the strains of LAB used were not capable of utilizing complex HMOs but metabolized HMO components and GOSs.

  6. Homogeneous human complex-type oligosaccharides in correctly folded intact glycoproteins: evaluation of oligosaccharide influence on protein folding, stability, and conformational properties.

    PubMed

    Kajihara, Yasuhiro; Tanabe, Yasutaka; Sasaoka, Shun; Okamoto, Ryo

    2012-05-07

    The N-glycosylation of proteins is generated at the consensus sequence NXS/T (where X is any amino acid except proline) by the biosynthetic process, and occurs in the endoplasmic reticulum and Golgi apparatus. In order to investigate the influence of human complex-type oligosaccharides on counterpart protein conformation, crambin and ovomucoide, which consist of 46 and 56 amino acid residues, respectively, were selected for synthesis of model glycoproteins. These small glycoproteins were intentionally designed to be glycosylated at the α-helix (crambin: 8 position), β-sheet (crambin: 2 position) and loop position between the antiparallel β-sheets (ovomucoide: 28 position), and were synthesized by using a peptide-segment coupling strategy. After preparation of these glycosylated polypeptide chains, protein folding experiments were performed under redox conditions by using cysteine-cystine. Although the small glycoproteins bearing intentional glycosylation at the α-helix and β-sheet exhibited a suitable folding process, glycosylation at the loop position between the antiparallel β-strands caused multiple products. The conformational differences in the isolated homogeneous glycoproteins compared with non-glycosylated counterparts were evaluated by circular dichroism (CD) and NMR spectroscopy. These analyses suggested that this intentional N-glycosylation did not result in large conformational changes in the purified protein structures, including the case of glycosylation at the loop position between the antiparallel β-strands. In addition to these experiments, the conformational properties of three glycoproteins were evaluated by CD spectroscopy under different temperatures. The oligosaccharides on the protein surface fluctuated considerably; this was dependent on the increase in the solution temperature and was thought to disrupt the protein tertiary structure. Based on the measurement of the CD spectra, however, the glycoproteins bearing three disulfide

  7. Efficient synthesis and activity of beneficial intestinal flora of two lactulose-derived oligosaccharides.

    PubMed

    Zhu, Zhen-Yuan; Cui, Di; Gao, Hui; Dong, Feng-Ying; Liu, Xiao-cui; Liu, Fei; Chen, Lu; Zhang, Yong-min

    2016-05-23

    Lactulose is considered as a prebiotic because it promotes the intestinal proliferation of Lactobacillus acidophilus which is added to various milk products. Moreover, lactulose is used in pharmaceuticals as a gentle laxative and to treat hyperammonemia. This study was aimed at the total synthesis of two Lactulose-derived oligosaccharides: one is 3-O-β-d-galactopyranosyl-d-fructose, d-fructose and β-d-galactose bounded together with β-1,3-glycosidic bound, the other is 1-O-β-d-galactopyranosyl-d-fructose, d-fructose and β-d-galactose bounded together with β-1,1-glycosidic bound, which were accomplished in seven steps from d-fructose and β-d-galactose and every step of yield above 75%. This synthetic route provided a practical and effective synthetic strategy for galactooligosaccharides, starting from commercially available monosaccharides. Then we evaluated on their prebiotic properties in the search for potential agents of regulating and improving the intestinal flora of human. The result showed that the prebiotic properties of Lactulose-derived oligosaccharides was much better than Lactulose. Among them, 3-O-β-d-galactopyranosyl-d-fructose displayed the most potent activity of proliferation of L. acidophilus.

  8. Extraction of green labeled pectins and pectic oligosaccharides from plant byproducts.

    PubMed

    Zykwinska, Agata; Boiffard, Marie-Hélène; Kontkanen, Hanna; Buchert, Johanna; Thibault, Jean-François; Bonnin, Estelle

    2008-10-08

    Green labeled pectins were extracted by an environmentally friendly way using proteases and cellulases being able to act on proteins and cellulose present in cell walls. Pectins were isolated from different plant byproducts, i.e., chicory roots, citrus peel, cauliflower florets and leaves, endive, and sugar beet pulps. Enzymatic extraction was performed at 50 degrees C for 4 h, in order to fulfill the conditions required for microbiological safety of extracted products. High methoxy (HM) pectins of high molar mass were extracted with three different enzyme mixtures. These pectins were subsequently demethylated with two pectin methyl esterases (PMEs), either the fungal PME from Aspergillus aculeatus or the orange PME. It was further demonstrated that high molar mass low methoxy (LM) pectins could also be extracted directly from cell walls by adding the fungal PME to the mixture of protease and cellulase. Moreover, health benefit pectic oligosaccharides, the so-called modified hairy regions, were obtained after enzymatic treatment of the residue recovered after pectin extraction. The enzymatic method demonstrates that it is possible to convert vegetable byproducts into high-added value compounds, such as pectins and pectic oligosaccharides, and thus considerably reduce the amount of these residues generated by food industries.

  9. Cranberry (Vaccinium macrocarpon) oligosaccharides decrease biofilm formation by uropathogenic Escherichia coli.

    PubMed

    Sun, Jiadong; Marais, Jannie P J; Khoo, Christina; LaPlante, Kerry; Vejborg, Rebecca M; Givskov, Michael; Tolker-Nielsen, Tim; Seeram, Navindra P; Rowley, David C

    2015-08-01

    The preventive effects of the American cranberry (Vaccinium macrocarpon) against urinary tract infections are supported by extensive studies which have primarily focused on its phenolic constituents. Herein, a phenolic-free carbohydrate fraction (designated cranf1b-F2) was purified from cranberry fruit using ion exchange and size exclusion chromatography. MALDI-TOF-MS analysis revealed that the cranf1b-F2 constituents are predominantly oligosaccharides possessing various degrees of polymerisation and further structural analysis (by GC-MS and NMR) revealed mainly xyloglucan and arabinan residues. In antimicrobial assays, cranf1b-F2 (at 1.25 mg/mL concentration) reduced biofilm production by the uropathogenic Escherichia coli CFT073 strain by over 50% but did not inhibit bacterial growth. Cranf1b-F2 (ranging from 0.625 - 10 mg/mL) also inhibited biofilm formation of the non-pathogenic E. coli MG1655 strain up to 60% in a concentration-dependent manner. These results suggest that cranberry oligosaccharides, in addition to its phenolic constituents, may play a role in its preventive effects against urinary tract infections.

  10. Cranberry (Vaccinium macrocarpon) oligosaccharides decrease biofilm formation by uropathogenic Escherichia coli

    PubMed Central

    Sun, Jiadong; Marais, Jannie P. J.; Khoo, Christina; LaPlante, Kerry; Vejborg, Rebecca M.; Givskov, Michael; Tolker-Nielsen, Tim; Seeram, Navindra P.; Rowley, David C.

    2015-01-01

    The preventive effects of the American cranberry (Vaccinium macrocarpon) against urinary tract infections are supported by extensive studies which have primarily focused on its phenolic constituents. Herein, a phenolic-free carbohydrate fraction (designated cranf1b-F2) was purified from cranberry fruit using ion exchange and size exclusion chromatography. MALDI-TOF-MS analysis revealed that the cranf1b-F2 constituents are predominantly oligosaccharides possessing various degrees of polymerisation and further structural analysis (by GC-MS and NMR) revealed mainly xyloglucan and arabinan residues. In antimicrobial assays, cranf1b-F2 (at 1.25 mg/mL concentration) reduced biofilm production by the uropathogenic Escherichia coli CFT073 strain by over 50% but did not inhibit bacterial growth. Cranf1b-F2 (ranging from 0.625 - 10 mg/mL) also inhibited biofilm formation of the non-pathogenic E. coli MG1655 strain up to 60% in a concentration-dependent manner. These results suggest that cranberry oligosaccharides, in addition to its phenolic constituents, may play a role in its preventive effects against urinary tract infections. PMID:26613004

  11. Label-Free Absolute Quantitation of Oligosaccharides Using Multiple Reaction Monitoring

    PubMed Central

    2015-01-01

    An absolute quantitation method for measuring free human milk oligosaccharides (HMOs) in milk samples was developed using multiple reaction monitoring (MRM). To obtain the best sensitivity, the instrument conditions were optimized to reduce the source and postsource fragmentation prior to the quadrupole transmission. Fragmentation spectra of HMOs using collision-induced dissociation were studied to obtain the best characteristic fragments. At least two MRM transitions were used to quantify and identify each structure in the same run. The fragment ions corresponded to the production of singly charged mono-, di-, and trisaccharide fragments. The sensitivity and accuracy of the quantitation using MRM were determined, with the detection limit in the femtomole level and the calibration range spanning over 5 orders of magnitude. Seven commercial HMO standards were used to create calibration curves and were used to determine a universal response for all HMOs. The universal response factor was used to estimate absolute amounts of other structures and the total oligosaccharide content in milk. The quantitation method was applied to 20 human milk samples to determine the variations in HMO concentrations from women classified as secretors and nonsecretors, a phenotype that can be identified by the concentration of 2′-fucosylation in their milk. PMID:24502421

  12. Synthesis, separation, and characterization of amphiphilic sulfated oligosaccharides enabled by reversed-phase ion pairing LC and LC-MS methods.

    PubMed

    Fenner, Amanda M; Kerns, Robert J

    2011-12-13

    Synthesis of amphiphilic oligosaccharides is problematic because traditional methods for separating and purifying oligosaccharides, including sulfated oligosaccharides, are generally not applicable to working with amphiphilic sugars. We report here RPIP-LC and LC-MS methods that enable the synthesis, separation, and characterization of amphiphilic N-arylacyl O-sulfonated aminoglycosides, which are being pursued as small-molecule glycosaminoglycan mimics. The methods described in this work for separating and characterizing these amphiphilic saccharides are further applied to a number of uses: monitoring the progression of sulfonation reactions with analytical RP-HPLC, characterizing sulfate content for individual molecules with ESI-MS, determining the degree of sulfation for products having mixed degrees of sulfation with HPLC and LC-MS, and purifying products with benchtop C18 column chromatography. We believe that the methods described here will be broadly applicable to enabling the synthesis, separation, and characterization of amphiphilic, sulfated, and phosphorylated oligosaccharides and other types of molecules substituted to varying degrees with both anionic and hydrophobic groups.

  13. Human milk oligosaccharides: evolution, structures and bioselectivity as substrates for intestinal bacteria.

    PubMed

    German, J Bruce; Freeman, Samara L; Lebrilla, Carlito B; Mills, David A

    2008-01-01

    Human milk contains a high concentration of diverse soluble oligosaccharides, carbohydrate polymers formed from a small number of monosaccharides. Novel methods combining liquid chromatography with high resolution mass spectrometry have identified approximately 200 unique oligosaccharides structures varying from 3 to 22 sugars. The increasing complexity of oligosaccharides follows the general pattern of mammalian evolution though the concentration and diversity of these structures in homo sapiens are strikingly. There is also diversity among human mothers in oligosaccharides. Milks from randomly selected mothers contain as few as 23 and as many as 130 different oligosaccharides. The functional implications of this diversity are not known. Despite the role of milk to serve as a sole nutrient source for mammalian infants, the oligosaccharides in milk are not digestible by human infants. This apparent paradox raises questions about the functions of these oligosaccharides and how their diverse molecular structures affect their functions. The nutritional function most attributed to milk oligosaccharides is to serve as prebiotics - a form of indigestible carbohydrate that is selectively fermented by desirable gut microflora. This function was tested by purifying human milk oligosaccharides and providing these as the sole carbon source to various intestinal bacteria. Indeed, the selectively of providing the complex mixture of oligosaccharides pooled from human milk samples is remarkable. Among a variety of Bifidobacteria tested only Bifidobacteria longum biovar infantis was able to grow extensively on human milk oligosaccharides as sole carbon source. The genomic sequence of this strain revealed approximately 700 genes that are unique to infantis, including a variety of co-regulated glycosidases, relative to other Bifidobacteria, implying a co-evolution of human milk oligosaccharides and the genetic capability of select intestinal bacteria to utilize them. The goal of

  14. Comparison of oligosaccharides in milk specimens from humans and twelve other species.

    PubMed

    Warren, C D; Chaturvedi, P; Newburg, A R; Oftedal, O T; Tilden, C D; Newburg, D S

    2001-01-01

    Human milk contains large amounts of many oligosaccharides, most of which are fucosylated; several inhibit pathogenic bacteria, viruses, and toxins that cause disease in humans. Although bovine milk is known to have much less and many fewer types of oligosaccharides, no studies heretofore have indicated whether the amount or complexity of human milk oligosaccharides is unique to our species. Toward this end, a comparison was made of the major individual oligosaccharides in milk specimens from a variety of species, including the great apes. The neutral compounds, which represent the bulk of oligosaccharides in human milk, were isolated, perbenzoylated, resolved by high performance liquid chromatography (HPLC), and detected at 229nm. Ambiguous structures were determined by mass spectrometry. All milk specimens contained lactose, although levels were quite low in bear and kangaroo milk. The types of oligosaccharides in milk specimens from the primates resembled those of human milk, but the amounts, especially of the larger molecules, were markedly lower. The relative amounts of oligosaccharides in the bonobo changed over the course of lactation, as they do in humans. Marine mammals generally had few oligosaccharides in their milk other than 2'-fucosyllactose. Grizzly and black bear milk specimens contained a wide range of oligosaccharides, many of which had novel, fucosylated structures. Milk specimens from humans, bears, and marsupials had the greatest quantity of, and the most complex, neutral oligosaccharides. Although human milk contained more oligosaccharide than did milk specimens from the other species studied, the presence of appreciable amounts of complex oligosaccharides was not unique to humans. This finding suggests that in animal milk specimens, as in human milk, neutral fucosylated oligosaccharides potentially offer protection from pathogens to offspring with immature immune systems.

  15. 1-N-glycyl beta-oligosaccharide derivatives as stable intermediates for the formation of glycoconjugate probes.

    PubMed

    Manger, I D; Rademacher, T W; Dwek, R A

    1992-11-10

    Incubation of reducing sugars in ammonium bicarbonate was found to be a simple procedure for the formation of beta-D-glycosylamines of purified complex oligosaccharides in 70-80% yield. These provide valuable intermediates for the synthesis of a wide range of oligosaccharide probes and derivatives by acylation of the 1-amino function. The 1-amino function showed different rates of reactivity with different reagents. In general, interactions with large ring systems such as the fluorophores dansyl chloride and carboxyfluorescein gave 10-20% yields of products, which consisted of mixtures of both anomeric forms, whereas smaller acylating reagents gave near-quantitative yields of the desired beta-D-derivatives. Steric effects may explain differences in reactivity. N-Chloroacetamido derivatives could be obtained in high yield with retention of the beta-anomeric configuration. Subsequent ammonolysis of the chloroacetamido function afforded the corresponding N-glycyl beta-derivatives. The linker thereby introduced retains the amino function, possesses the useful properties of fixed anomeric configuration, improved stability, and uniform reactivity with a variety of reagents, and is structurally analogous to an asparagine side chain. The potential therefore exists for the generation of oligosaccharide derivatives tailored for different applications.

  16. Evaluation of oligosaccharide synthesis from lactose and lactulose using β-galactosidases from Kluyveromyces isolated from artisanal cheeses.

    PubMed

    Padilla, Beatriz; Ruiz-Matute, Ana I; Belloch, Carmela; Cardelle-Cobas, Alejandra; Corzo, Nieves; Manzanares, Paloma

    2012-05-23

    The β-galactosidase activity of 15 Kluyveromyces strains isolated from cheese belonging to Kluyveromyces lactis and Kluyveromyces marxianus species was tested for the production of oligosaccharides derived from lactose (GOS) and lactulose (OsLu). All Kluyveromyces crude cell extracts (CEEs) produced GOS, such as 6-galactobiose and 3'-, 4'-, and 6'-galactosyl-lactose. At 4 h of reaction, the main trisaccharide formed was 6'-galactosyl-lactose (20 g/100 g of total carbohydrates). The formation of OsLu was also observed by all CEEs tested, with 6-galactobiose, 6'-galactosyl-lactulose, and 1-galactosyl-lactulose being found in all of the reaction mixtures. The synthesis of trisaccharides predominated over other oligosaccharides. K. marxianus strain O3 produced the highest yields of GOS and OsLu after 4 h of reaction, reaching 42 g/100 g of total carbohydrates (corresponding to 80% lactose hydrolysis) and 45 g/100 g of total carbohydrates (corresponding to 87% lactulose hydrolysis), respectively. Therefore, the present study contributes to a better insight into dairy Kluyveromyces β-galactosidases and shows the feasibility of these enzymes to transglycosylate lactose and lactulose, producing high yields of prebiotic oligosaccharides.

  17. Mrt, a gene unique to fungi, encodes an oligosaccharide transporter and facilitates rhizosphere competency in Metarhizium robertsii.

    PubMed

    Fang, Weiguo; St Leger, Raymond J

    2010-11-01

    The symbiotic associations between rhizospheric fungi and plants have enormous environmental impact. Fungi are crucial to plant health as antagonists of pathogens and herbivores and facilitate the uptake of soil nutrients. However, little is known about the plant products obtained by fungi in exchange or how they are transported through the symbiotic interface. Here, we demonstrate that sucrose and raffinose family oligosaccharides in root exudates are important for rhizosphere competence in the insect pathogen Metarhizium robertsii (formerly known as Metarhizium anisopliae). We identified mutants in the Metarhizium raffinose transporter (Mrt) gene of M. robertsii that grew poorly in root exudate and were greatly reduced in rhizosphere competence on grass roots. Studies on sugar uptake, including competition assays, revealed that MRT was a sucrose and galactoside transporter. Disrupting MRT resulted in greatly reduced or no growth on sucrose and galactosides but did not affect growth on monosaccharides or oligosaccharides composed entirely of glucose subunits. Consistent with this, expression of Mrt is exclusively up-regulated by galactosides and sucrose. Expressing a green fluorescent protein gene under the control of the Mrt promoter confirmed that MRT was expressed by germlings in the vicinity of grass roots but not in surrounding bulk soil. Disrupting Mrt did not reduce virulence to insects, demonstrating that Mrt is exclusively involved in M. robertsii's interactions with plants. To our knowledge, MRT is the first oligosaccharide transporter identified and characterized in a fungus and is unique to filamentous fungi, but homologous genes in Magnaporthe, Ustilago, Aspergillus, Fusarium, Epichloe, and Penicillium species indicate that oligosaccharide transport is of widespread significance.

  18. Specific lignin precipitation for oligosaccharides recovery from hot water wood extract.

    PubMed

    Chen, Xiaoqian; Wang, Zhaojiang; Fu, Yingjuan; Li, Zongquan; Qin, Menghua

    2014-01-01

    Hot water extraction is an important strategy of wood fractionation, by which the hemicelluloses can be separated for value-added products, while the residual solid can still be processed into traditional wood products. In this study, a combined process consisting of specific lignin precipitation and dialysis was proposed to recover hemicellulosic oligosaccharides (OS) from hot water extract (HWE). The results showed that polyaluminium chloride (PAC) precipitation was highly specific to large molecular lignin, leading to 25.1% lignin removal with negligible OS loss through charge neutralization mechanism. The separation was further enhanced by dialysis, reaching 37.6% OS recovery from HWE with remarkable purity of 94.1%. By the proposed process, 56.36 g OS, mainly xylooligosaccharides with two fractions of 5.2 and 0.51 kDa was recovered from one kg dried wood. This process can be envisaged as a great contribution to wood biorefinery. Copyright © 2013 Elsevier Ltd. All rights reserved.

  19. Release of feruloylated oligosaccharides from wheat bran through submerged fermentation by edible mushrooms.

    PubMed

    Xie, Chunyan; Wu, Zhiyan; Guo, Hongzhen; Gu, Zhenxin

    2014-07-01

    Wheat bran, a by-product of the flour industry, is believed to be a raw material for the production of feruloylated oligosaccharides (FOs) because of its high content of conjiont ferulic acid (FA). Studies were carried out to identify edible mushrooms that are able to release FOs from wheat bran. All the six tested mushrooms (Pleurotus ostreatus, Hericium erinaceum, Auricularia auricula, Cordyceps militaris, Agrocybe chaxingu, and Ganoderma lucium) were found to release FOs, and Agrocybe chaxingu had the highest yield, reaching 35.4 µM in wheat bran broth. Enzymes detection showed that these species secreted extracellular enzymes during fermentation, including cellulase and xylanase. Agrocybe chaxingu secreted the significant amount of xylanase (180 mU ml(-1) ), which was responsible for the release of FOs from wheat bran, while Hericium erinaceum secreted FA esterase which could disassemble FOs.

  20. Capillary electrophoresis of neutral carbohydrates: mono-, oligosaccharides, glycosides.

    PubMed

    Campa, Cristiana; Rossi, Marco

    2008-01-01

    This chapter reports an overview of the recent advances in the analysis of neutral sugars by capillary electrophoresis (CE); furthermore, some relevant reviews and research articles in the field are tabulated. Comparison of CE with chromatography is also presented, with special attention to separation efficiency and sensitivity. The main routes aimed at pretreatment and CE analysis of uncharged mono-, oligosaccharides, and glycosides are described. Representative examples of such procedures are reported in detail, upon describing robust methodologies for the study of (1) neutral mono- and oligosaccharides derivatized by reductive amination and by formation of glycosylamines; (2) underivatized mono- and di-saccharides analyzed using highly alkaline buffers; and (3) anomeric couples of glycosides separated using borate-based buffers.

  1. Nursing our microbiota: molecular linkages between bifidobacteria and milk oligosaccharides

    PubMed Central

    Sela, David A.; Mills, David A.

    2010-01-01

    As the sole nutrition provided to infants, bioactive molecules dissolved in milk influence the development of our gut microbiota. Accordingly, human milk oligosaccharides (HMOs) are minimally digested by the infant and persist to negatively and positively regulate gut microbiota. Accordingly, infant-type bifidobacteria utilize these soluble carbohydrate oligomers by convergent mechanisms. Bifidobacterium longum subsp. infantis efficiently consumes several small mass HMOs and possesses a large gene cluster and other loci dedicated to HMO metabolism. In contrast, adult-associated bifidobacteria such as the closely related B. longum subsp. longum, are deficient for HMO utilization, although they retain the capacity to ferment plant oligosaccharides and constituent pentose sugars. Thus, the ability to subsist on HMO may demark infant-associated ecotypes as these bifidobacteria may have adapted to colonize the nursing infant. PMID:20409714

  2. Inulin, a flexible oligosaccharide. II: Review of its pharmaceutical applications.

    PubMed

    Mensink, Maarten A; Frijlink, Henderik W; van der Voort Maarschalk, Kees; Hinrichs, Wouter L J

    2015-12-10

    Inulin is a flexible oligosaccharide which has been used primarily in food for decades. Recently new applications in the pharmaceutical arena were described. In a previous review (Mensink et al. (2015). Carbohydrate Polymers, 130, 405) we described the physicochemical characteristics of inulin, characteristics which make inulin a highly versatile substance. Here, we review its pharmaceutical applications. Applications of inulin that are addressed are stabilization of proteins, modified drug delivery (dissolution rate enhancement and drug targeting), and lastly physiological and disease-modifying effects of inulin. Further uses of inulin include colon specific drug administration and stabilizing and adjuvating vaccine formulations. Overall, the uses of inulin in the pharmaceutical area are very diverse and research is still continuing, particularly with chemically modified inulins. It is therefore likely that even more applications will be found for this flexible oligosaccharide.

  3. Advances in Analysis of Human Milk Oligosaccharides123

    PubMed Central

    Ruhaak, L. Renee; Lebrilla, Carlito B.

    2012-01-01

    Oligosaccharides in human milk strongly influence the composition of the gut microflora of neonates. Because it is now clear that the microflora play important roles in the development of the infant immune system, human milk oligosaccharides (HMO) are studied frequently. Milk samples contain complex mixtures of HMO, usually comprising several isomeric structures that can be either linear or branched. Traditionally, HMO profiling was performed using HPLC with fluorescence or UV detection. By using porous graphitic carbon liquid chromatography MS, it is now possible to separate and identify most of the isomers, facilitating linkage-specific analysis. Matrix-assisted laser desorption ionization time-of-flight analysis allows fast profiling, but does not allow isomer separation. Novel MS fragmentation techniques have facilitated structural characterization of HMO that are present at lower concentrations. These techniques now facilitate more accurate studies of HMO consumption as well as Lewis blood group determinations. PMID:22585919

  4. Structure-function relationships of human milk oligosaccharides.

    PubMed

    Bode, Lars; Jantscher-Krenn, Evelyn

    2012-05-01

    Human milk contains more than a hundred structurally distinct oligosaccharides. In this review, we provide examples of how the structural characteristics of these human milk oligosaccharides (HMO) determine functionality. Specific α1-2-fucosylated HMO have been shown to serve as antiadhesive antimicrobials to protect the breast-fed infant against infections with Campylobacter jejuni, one of the most common causes of bacterial diarrhea. In contrast, α1-2-fucosylation may abolish the beneficial effects of HMO against Entamoeba histolytica, a protozoan parasite that causes colitis, acute dysentery, or chronic diarrhea. In a different context, HMO need to be both fucosylated and sialylated to reduce selectin-mediated leukocyte rolling, adhesion, and activation, which may protect breast-fed infants from excessive immune responses. In addition, our most recent data show that a single HMO that carries not 1 but 2 sialic acids protects neonatal rats from necrotizing enterocolitis, one of the most common and often fatal intestinal disorders in preterm infants. Oligosaccharides currently added to infant formula are structurally different from the oligosaccharides naturally occurring in human milk. Thus, it appears unlikely that they can mimic some of the structure-specific effects of HMO. Recent advances in glycan synthesis and isolation have increased the availability of certain HMO tri- and tetrasaccharides for in vitro and in vivo preclinical studies. In the end, intervention studies are needed to confirm that the structure-specific effects observed at the laboratory bench translate into benefits for the human infant. Ultimately, breastfeeding remains the number one choice to nourish and nurture our infants.

  5. Characterization of protein-linked oligosaccharides in trypanosomatid flagellates.

    PubMed

    Mendelzon, D H; Previato, J O; Parodi, A J

    1986-03-01

    Protein-linked, endo-beta-N-acetylglucosaminidase H-sensitive oligosaccharides were isolated from several trypanosomatids incubated with [U-14C]glucose. Structural analysis of the compounds revealed that Man9GlcNAc2 was the oligosaccharide transferred from dolichol-P-P derivatives to proteins in Trypanosoma dionisii, Trypanosoma conorhini, Leptomonas samueli and Herpetomonas samuelpessoai and Man6GlcNAc2 in Blastocrithidia culicis and Leishmania adleri. In all cases, transiently glucosylated compounds were detected: Glc1Man7-9GlcNAc2 in T. dionisii, T. conorhini, L. samueli; Glc1Man9GlcNAc2 in H. samuelpessoai, Glc1Man6GlcNAc2 in B. culicis and Glc1Man6GlcNAc2 and Glc1Man5GlcNAc2 in L. adleri. The mechanism of protein glycosylation in T. dionisii and T. conorhini appeared to be similar to that described before for Trypanosoma cruzi epimastigotes, although some differences were found between the structures of the main isomers of Man7GlcNAc2 and Man8GlcNAc2 present in T. conorhini and T. cruzi. Differences between the mechanisms of glycosylation occurring in Leishmania mexicana and L. adleri were also found: Man6GlcNAc2 in the latter microorganism was demannosylated to Man5GlcNAc2, a step not detected in the former parasite. A novel substituent in N-linked high mannose-type oligosaccharides was found in L. samueli and H. samuelpessoai: galactose in the furanose configuration. In the latter trypanosomatid, Man9GlcNAc2 was demannosylated only to Man8GlcNAc2, whereas in all other parasites in which the same oligosaccharide was transferred to proteins, Man5-7GlcNAc2 were also detected.

  6. Gliding Motility of Mycoplasma mobile on Uniform Oligosaccharides

    PubMed Central

    Kasai, Taishi; Hamaguchi, Tasuku

    2015-01-01

    ABSTRACT The binding and gliding of Mycoplasma mobile on a plastic plate covered by 53 uniform oligosaccharides were analyzed. Mycoplasmas bound to and glided on only 21 of the fixed sialylated oligosaccharides (SOs), showing that sialic acid is essential as the binding target. The affinities were mostly consistent with our previous results on the inhibitory effects of free SOs and suggested that M. mobile recognizes SOs from the nonreducing end with four continuous sites as follows. (i and ii) A sialic acid at the nonreducing end is tightly recognized by tandemly connected two sites. (iii) The third site is recognized by a loose groove that may be affected by branches. (iv) The fourth site is recognized by a large groove that may be enhanced by branches, especially those with a negative charge. The cells glided on uniform SOs in manners apparently similar to those of the gliding on mixed SOs. The gliding speed was related inversely to the mycoplasma's affinity for SO, suggesting that the detaching step may be one of the speed determinants. The cells glided faster and with smaller fluctuations on the uniform SOs than on the mixtures, suggesting that the drag caused by the variation in SOs influences gliding behaviors. IMPORTANCE Mycoplasma is a group of bacteria generally parasitic to animals and plants. Some Mycoplasma species form a protrusion at a pole, bind to solid surfaces, and glide in the direction of the protrusion. These procedures are essential for parasitism. Usually, mycoplasmas glide on mixed sialylated oligosaccharides (SOs) derived from glycoprotein and glycolipid. Since gliding motility on uniform oligosaccharides has never been observed, this study gives critical information about recognition and interaction between receptors and SOs. PMID:26148712

  7. Structure-Function Relationships of Human Milk Oligosaccharides123

    PubMed Central

    Bode, Lars; Jantscher-Krenn, Evelyn

    2012-01-01

    Human milk contains more than a hundred structurally distinct oligosaccharides. In this review, we provide examples of how the structural characteristics of these human milk oligosaccharides (HMO) determine functionality. Specific α1–2-fucosylated HMO have been shown to serve as antiadhesive antimicrobials to protect the breast-fed infant against infections with Campylobacter jejuni, one of the most common causes of bacterial diarrhea. In contrast, α1–2-fucosylation may abolish the beneficial effects of HMO against Entamoeba histolytica, a protozoan parasite that causes colitis, acute dysentery, or chronic diarrhea. In a different context, HMO need to be both fucosylated and sialylated to reduce selectin-mediated leukocyte rolling, adhesion, and activation, which may protect breast-fed infants from excessive immune responses. In addition, our most recent data show that a single HMO that carries not 1 but 2 sialic acids protects neonatal rats from necrotizing enterocolitis, one of the most common and often fatal intestinal disorders in preterm infants. Oligosaccharides currently added to infant formula are structurally different from the oligosaccharides naturally occurring in human milk. Thus, it appears unlikely that they can mimic some of the structure-specific effects of HMO. Recent advances in glycan synthesis and isolation have increased the availability of certain HMO tri- and tetrasaccharides for in vitro and in vivo preclinical studies. In the end, intervention studies are needed to confirm that the structure-specific effects observed at the laboratory bench translate into benefits for the human infant. Ultimately, breastfeeding remains the number one choice to nourish and nurture our infants. PMID:22585916

  8. The chromatographic analysis of oligosaccharides and preparation of 1-kestose and nystose in yacon.

    PubMed

    Zhu, Zhen-Yuan; Lian, Hong-Yu; Si, Chuan-Ling; Liu, Yang; Liu, Nian; Chen, Jing; Ding, Li-Na; Yao, Qiang; Zhang, Yongmin

    2012-05-01

    The thin-layer chromatographic analysis of the crude oligosaccharides extracted from yacon revealed the presence of glucose, fructose, sucrose, 1-kestose and nystose. The qualitative and quantitative analysis was carried out on oligosaccharides by high pressure liquid chromatography and the results showed that the contents of d-glucose, fructose, sucrose, 1-kestose, nystose and 1-fructofuranosyl nystose in oligosaccharides were 38.30%, 16.44%, 14.58%, 12.29%, 12.17%, 6.20%, respectively. The content of the fructooligosaccharides in oligosaccharides was 30.66%. The crude oligosaccharides were separated and purified by silica gel column chromatography. The two fractions obtained from crude oligosaccharides were 1-kestose and nystose, which were identified by mass spectra. The yield of 1-kestose and nystose were 10.36% and 9.73%, respectively. The purity of 1-kestose was 82.9% and of nystose was 73.6%.

  9. In vitro fermentability of sugar beet pulp derived oligosaccharides using human and pig fecal inocula.

    PubMed

    Leijdekkers, Antonius G M; Aguirre, Marisol; Venema, Koen; Bosch, Guido; Gruppen, Harry; Schols, Henk A

    2014-02-05

    The in vitro fermentation characteristics of different classes of sugar beet pectic oligosaccharides (SBPOS) were studied using human and pig fecal inocula. The SBPOS consisted mainly of partially acetylated rhamnogalacturonan oligosaccharides and partially methyl-esterified/acetylated homogalacturonan oligosaccharides. Some SBPOS contained an unsaturated galacturonic acid residue at their non-reducing end. It was shown that SBPOS could be completely fermented by human and pig fecal microbiota, thereby producing butyrate yet mainly acetate and propionate as metabolites. The degradation of SBPOS by pig fecal microbiota was different and much slower compared to human fecal microbiota. In general, rhamnogalacturonan oligosaccharides were degraded slower than homogalacturonan oligosaccharides. Acetylation of rhamnogalacturonan oligosaccharides lowered the degradation rate by pig fecal microbiota but not by human fecal microbiota. No classic bifidogenic effect was shown for SBPOS using human fecal inoculum. However, several other potentially interesting modifications in the microbiota composition that can be associated with host health were observed, which are discussed.

  10. Novel Alginate Lyase (Aly5) from a Polysaccharide-Degrading Marine Bacterium, Flammeovirga sp. Strain MY04: Effects of Module Truncation on Biochemical Characteristics, Alginate Degradation Patterns, and Oligosaccharide-Yielding Properties

    PubMed Central

    Han, Wenjun; Gu, Jingyan; Cheng, Yuanyuan; Liu, Huihui; Li, Yuezhong

    2015-01-01

    Alginate lyases are important tools for oligosaccharide preparation, medical treatment, and energy bioconversion. Numerous alginate lyases have been elucidated. However, relatively little is known about their substrate degradation patterns and product-yielding properties, which is a limit to wider enzymatic applications and further enzyme improvements. Herein, we report the characterization and module truncation of Aly5, the first alginate lyase obtained from the polysaccharide-degrading bacterium Flammeovirga. Aly5 is a 566-amino-acid protein and belongs to a novel branch of the polysaccharide lyase 7 (PL7) superfamily. The protein rAly5 is an endolytic enzyme of alginate and associated oligosaccharides. It prefers guluronate (G) to mannuronate (M). Its smallest substrate is an unsaturated pentasaccharide, and its minimum product is an unsaturated disaccharide. The final alginate digests contain unsaturated oligosaccharides that generally range from disaccharides to heptasaccharides, with the tetrasaccharide fraction constituting the highest mass concentration. The disaccharide products are identified as ΔG units. While interestingly, the tri- and tetrasaccharide fractions each contain higher proportions of ΔG to ΔM ends, the larger final products contain only ΔM ends, which constitute a novel oligosaccharide-yielding property of guluronate lyases. The deletion of the noncatalytic region of Aly5 does not alter its M/G preference but significantly decreases the enzymatic activity and enzyme stability. Notably, the truncated protein accumulates large final oligosaccharide products but yields fewer small final products than Aly5, which are codetermined by its M/G preference to and size enlargement of degradable oligosaccharides. This study provides novel enzymatic properties and catalytic mechanisms of a guluronate lyase for potential uses and improvements. PMID:26519393

  11. Novel Alginate Lyase (Aly5) from a Polysaccharide-Degrading Marine Bacterium, Flammeovirga sp. Strain MY04: Effects of Module Truncation on Biochemical Characteristics, Alginate Degradation Patterns, and Oligosaccharide-Yielding Properties.

    PubMed

    Han, Wenjun; Gu, Jingyan; Cheng, Yuanyuan; Liu, Huihui; Li, Yuezhong; Li, Fuchuan

    2015-10-30

    Alginate lyases are important tools for oligosaccharide preparation, medical treatment, and energy bioconversion. Numerous alginate lyases have been elucidated. However, relatively little is known about their substrate degradation patterns and product-yielding properties, which is a limit to wider enzymatic applications and further enzyme improvements. Herein, we report the characterization and module truncation of Aly5, the first alginate lyase obtained from the polysaccharide-degrading bacterium Flammeovirga. Aly5 is a 566-amino-acid protein and belongs to a novel branch of the polysaccharide lyase 7 (PL7) superfamily. The protein rAly5 is an endolytic enzyme of alginate and associated oligosaccharides. It prefers guluronate (G) to mannuronate (M). Its smallest substrate is an unsaturated pentasaccharide, and its minimum product is an unsaturated disaccharide. The final alginate digests contain unsaturated oligosaccharides that generally range from disaccharides to heptasaccharides, with the tetrasaccharide fraction constituting the highest mass concentration. The disaccharide products are identified as ΔG units. While interestingly, the tri- and tetrasaccharide fractions each contain higher proportions of ΔG to ΔM ends, the larger final products contain only ΔM ends, which constitute a novel oligosaccharide-yielding property of guluronate lyases. The deletion of the noncatalytic region of Aly5 does not alter its M/G preference but significantly decreases the enzymatic activity and enzyme stability. Notably, the truncated protein accumulates large final oligosaccharide products but yields fewer small final products than Aly5, which are codetermined by its M/G preference to and size enlargement of degradable oligosaccharides. This study provides novel enzymatic properties and catalytic mechanisms of a guluronate lyase for potential uses and improvements.

  12. Separation of radiolabelled glycosaminoglycan oligosaccharides by polyacrylamide-gel electrophoresis.

    PubMed

    Hampson, I N; Gallagher, J T

    1984-08-01

    Glycosaminoglycan oligosaccharides generated by treatment of biosynthetically radiolabelled dermatan sulphate and hyaluronic acid with chondroitin AC lyase or testicular hyaluronidase may be resolved into a series of discrete bands by polyacrylamide-gel electrophoresis. Bands were identified by fixation in glacial acetic acid containing 20% (w/v) 2,5-diphenyloxazole followed by fluorography. The bands represented glycans which differed in size by one disaccharide unit. For the larger oligosaccharides (decasaccharides and above) of similar charge: mass ratio, there was a linear relationship between electrophoretic mobility and log Mr. However, the smaller species showed anomalous migration patterns. Consideration of the structures of the fragments produced by the different enzyme treatments suggests that copolymeric and homopolymeric oligosaccharides may be separated by polyacrylamide-gel electrophoresis. There are many potential applications of this technique, foremost amongst them being studies on the molecular size heterogeneity and patterns of enzyme-mediated depolymerization of native glycosaminoglycan chains and investigations into rates of polymer chain elongation and post-polymerization modification reactions so essential to glycosaminoglycan function.

  13. Annotation and Structural Analysis of Sialylated Human Milk Oligosaccharides

    PubMed Central

    Wu, Shuai; Grimm, Rudolf; German, J. Bruce; Lebrilla, Carlito B.

    2011-01-01

    Sialylated human milk oligosaccharides (SHMOs) are important components of human milk oligosaccharides. Sialic acids are typically found on the nonreducing end and are known binding sites for pathogens and aid in neonates’ brain development. Due to their negative charge and hydrophilic nature, they also help modulate cell-cell interactions. It has also been shown that sialic acids are involved in regulating the immune response and aid in brain development. In this study, the enriched SHMOs from pooled milk sample were analyzed by HPLC-Chip/QTOF MS. The instrument employs a microchip-based nano-LC column packed with porous graphitized carbon (PGC) to provide excellent isomer separation for SHMOs with highly reproducible retention time. The precursor ions were further examined with collision-induced dissociation (CID). By applying the proper collision energy, isomers can be readily differentiated by diagnostic peaks and characteristic fragmentation patterns. A set of 30 SHMO structures with retention times, accurate masses and MS/MS spectra was deduced and incorporated into an HMO library. When combined with previously determined neutral components, a library with over 70 structures is obtained allowing high-throughput oligosaccharide structure identification. PMID:21133381

  14. Synthesis of heparin-like oligosaccharides on polymer supports.

    PubMed

    Ojeda, Rafael; Terentí, Olimpia; de Paz, José-Luis; Martín-Lomas, Manuel

    2004-01-01

    The biological functions of a variety of proteins are regulated by heparan sulfate glycosaminoglycans. In order to facilitate the elucidation of the molecular basis of glycosaminoglycan-protein interactions we have developed syntheses of heparin-like oligosaccharides on polymer supports. A completely stereoselective strategy previously developed by us for the synthesis of these oligosaccharides in solution has been extended to the solid phase using an acceptor-bound approach. Both a soluble polymer support and a polyethylene glycol-grafted polystyrene resin have been used and different strategies for the attachment of the acceptor to the support have been explored. The attachment of fully protected disaccharide building blocks to a soluble support through the carboxylic group of the uronic acid unit by a succinic ester linkage, the use of trichloroacetimidates as glycosylating agents and of a functionalized Merryfield type resin for the capping process allowed for the construction of hexasaccharide and octasaccharide fragments containing the structural motif of the regular region of heparin. This strategy may facilitate the synthesis of glycosaminoglycan oligosaccharides by using the required building blocks in the glycosylation sequence.

  15. Engineering chemical reactivity on cell surfaces through oligosaccharide biosynthesis

    SciTech Connect

    Mahal, L.K.; Yareme, K.J.; Bertozzi, C.R.

    1997-05-16

    Cell surface oligosaccharide can be engineered to display unusual functional groups for the selective chemical remodeling of cell surfaces. An unnatural derivative of N-acetyl-mannosamine, which has a ketone group, was converted to the corresponding sialic acid and incorporated into cell surface oligosaccharide metabolically, resulting in the cell surface display of ketone groups. The ketone group on the cell surface can then be covalently ligated under physiological conditions with molecules carrying a complementary reactive functional group such as the hydrazide. Cell surface reactions of this kind should prove useful in the introduction of new recognition epitopes, such as peptides, oligosaccharide, or small organic molecules, onto cell surfaces and in the subsequent modulation of cell-cell or cell-small molecule binding events. The versatility of this technology was demonstrated by an example of selective drug delivery. Cells were decorated with biotin through selective conjugation to ketone groups, and selectively killed in the presence of a ricin A chain-avidin conjugate. 30 refs., 4 figs.

  16. Neoagaro-oligosaccharide monomers inhibit inflammation in LPS-stimulated macrophages through suppression of MAPK and NF-κB pathways

    PubMed Central

    Wang, Wei; Liu, Pei; Hao, Cui; Wu, Lijuan; Wan, Wenjin; Mao, Xiangzhao

    2017-01-01

    Neoagaro-oligosaccharides derived from agarose have been demonstrated to possess a variety of biological activities, such as anti-bacteria and anti-oxidative activities. In this study, we mainly explored the inhibitory effects and the mechanisms of neoagaro-oligosaccharide monomers against LPS-induced inflammatory responses in mouse macrophage RAW264.7 cells. The results indicated that neoagaro-oligosaccharide monomers especially neoagarotetraose could significantly reduce the production and release of NO in LPS-induced macrophages. Neoagarotetraose significantly suppressed the expression and secretion of inducible nitric oxide synthase (iNOS) and proinflammatory cytokines such as TNF-α and IL-6. The inhibition mechanisms may be associated with the inhibition of the activation of p38MAPK, Ras/MEK/ERK and NF-κB signaling pathways. Thus, neoagarotetraose may attenuate the inflammatory responses through downregulating the MAPK and NF-κB signaling pathways in LPS-stimulated macrophages. In summary, the marine-derived neoagaro-oligosaccharide monomers merit further investigation as novel anti-inflammation agents in the future. PMID:28266652

  17. 1H NMR characterization of a hen ovalbumin tyrosinamide N-linked oligosaccharide library.

    PubMed

    Corradi Da Silva, M L; Stubbs, H J; Tamura, T; Rice, K G

    1995-04-20

    A library of 15 N-linked oligosaccharide structures was prepared from ovalbumin and characterized using high-field NMR and mass spectrometry. The oligosaccharides were enzymatically released from ovalbumin glycopeptides, and the reducing ends were reacted with ammonium bicarbonate to form oligosaccharide-glycosylamines. These reacted with Boc-tyrosine-N-hydroxysuccinimide ester, resulting in a mixture of tyrosinamide-oligosaccharides. The Boc group was removed to expose an amine terminus which enhanced the resolution of tyrosinamide-oligosaccharides when chromatographed on reverse-phase HPLC. Ten major and five minor oligosaccharides were purified on a micromole scale and characterized using 1H NMR and FAB-MS. The structures include high-mannose, hybrid, and complex oligosaccharides possessing from two to five antenna, providing the most complete definition of ovalbumin N-linked oligosaccharides to date. The resulting library is well suited to biological studies due to the presence of a single terminal tyrosine residue on each oligosaccharide that allows radioiodination or the attachment of additional probes to these glycoconjugates prior to biological studies.

  18. Structural study of asparagine-linked oligosaccharide moiety of taste-modifying protein, miraculin.

    PubMed

    Takahashi, N; Hitotsuya, H; Hanzawa, H; Arata, Y; Kurihara, Y

    1990-05-15

    The structures of the N-linked oligosaccharides of miraculin, which is a taste modifying glycoprotein isolated from miracle fruits, berries of Richadella dulcifica, are reported. Asparagine-linked oligosaccharides were released from the protein by glycopeptidase (almond) digestion. The reducing ends of the oligosaccharide chains thus obtained were aminated with a fluorescent reagent, 2-aminopyridine, and the mixture of pyridylamino derivatives of the oligosaccharides was separated by high performance liquid chromatography (HPLC) on an ODS-silica column. More than five kinds of oligosaccharide fractions were separated by the one chromatographic run. The structure of each oligosaccharide thus isolated was analyzed by a combination of sequential exoglycosidase digestion and another kind of HPLC with an amidesilica column. Furthermore, high resolution proton nuclear magnetic resonance (1H NMR) measurements were carried out. It was found that 1) five oligosaccharides obtained are a series of compounds with xylose-containing common structural core, Xyl beta 1----2 (Man alpha 1----6) Man beta 1----4-GlcNAc beta 1----4 (Fuca1----3)GlcNAc, 2) a variety of oligosaccharide structures are significant for two glycosylation sites, Asn-42 and Asn-186, and 3) two new oligosaccharides, B and D, with unusual structures containing monoantennary complex-type were characterized. (formula; see text)

  19. A new sialyloligosaccharide from human milk: isolation and characterization using anti-oligosaccharide antibodies

    SciTech Connect

    Prieto, P.A.; Smith, D.F.

    1984-03-01

    A previously undescribed sialyloligosaccharide has been isolated from human milk using a specific anti-sialyloligosaccharide antibody. Structural studies of the radiolabeled oligosaccharide by enzyme degradation and binding by specific anti-oligosaccharide sera are consistent with the following structure: (sequence in text) The oligosaccharide is present only in milk from donors who secrete A, B, or H blood group substances; this is consistent with the requirement of at least one copy of the Se (Secretor) gene necessary for the synthesis of oligosaccharides with Fuc alpha 1-2Gal . . . linkages.

  20. An Update on Oligosaccharides and Their Esters from Traditional Chinese Medicines: Chemical Structures and Biological Activities

    PubMed Central

    Chen, Xiang-Yang; Wang, Ru-Feng; Liu, Bin

    2015-01-01

    A great number of naturally occurring oligosaccharides and oligosaccharide esters have been isolated from traditional Chinese medicinal plants, which are used widely in Asia and show prominent curative effects in the prevention and treatment of kinds of diseases. Numerous in vitro and in vivo experiments have revealed that oligosaccharides and their esters exhibited various activities, including antioxidant, antidepressant, cytotoxic, antineoplastic, anti-inflammatory, neuroprotective, cerebral protective, antidiabetic, plant growth-regulatory, and immunopotentiating activities. This review summarizes the investigations on the distribution, chemical structures, and bioactivities of natural oligosaccharides and their esters from traditional Chinese medicines between 2003 and 2013. PMID:25861364

  1. Chemical characterization of milk oligosaccharides of the koala (Phascolarctos cinereus).

    PubMed

    Urashima, Tadasu; Taufik, Epi; Fukuda, Rino; Nakamura, Tadashi; Fukuda, Kenji; Saito, Tadao; Messer, Michael

    2013-11-01

    Previous structural characterizations of marsupial milk oligosaccharides had been performed in only two macropod species, the tammar wallaby and the red kangaroo. To clarify the homology and heterogeneity of milk oligosaccharides among marsupial species, which could provide information on their evolution, the oligosaccharides of the koala milk carbohydrate fraction were characterized in this study. Neutral and acidic oligosaccharides were separated from the carbohydrate fraction of milk of the koala, a non-macropod marsupial, and characterized by (1)H-nuclear magnetic resonance spectroscopy. The structures of the neutral saccharides were found to be Gal(β1-4)Glc (lactose), Gal(β1-3)Gal(β1-4)Glc (3'-galactosyllactose), Gal(β1-3)Gal(β1-3)Gal(β1-4)Glc (3',3″-digalactosyllactose), Gal(β1-3)[Gal(β1-4)GlcNAc(β1-6)]Gal(β1-4)Glc (lacto-N-novopentaose I) and Gal(β1-3){Gal(β1-4)[Fuc(α1-3)]GlcNAc(β1-6)}Gal(β1-4)Glc (fucosyl lacto-N-novopentaose I), while those of the acidic saccharides were Neu5Ac(α2-3)Gal(β1-4)Glc (3'-SL), Neu5Ac(α2-3)Gal(β1-3)Gal(β1-4)Gal (sialyl 3'-galactosyllactose), Neu5Ac(α2-3)Gal(β1-3)[Gal(β1-4)GlcNAc(β1-6)]Gal(β1-4)Glc (sialyl lacto-N-novopentaose a), Gal(β1-3)[Neu5Ac(α2-6)Gal(β1-4)GlcNAc(β1-6)]Gal(β1-4)Glc (sialyl lacto-N-novopentaose b), Gal(β1-3)[Neu5Ac(α2-3)Gal(β1-4)GlcNAc(β1-6)]Gal(β1-4)Glc (sialyl lacto-N-novopentaose c), and Neu5Ac(α2-3)Gal(β1-3){Gal(β1-4)[Fuc(α1-3)]GlcNAc(β1-6)}Gal(β1-4)Glc (fucosyl sialyl lacto-N-novopentaose a). The neutral oligosaccharides, other than fucosyl lacto-N-novopentaose I, a novel hexasaccharide, had been found in milk of the tammar wallaby, a macropod marsupial, while the acidic oligosaccharides, other than fucosyl sialyl lacto-N-novopentaose a had been identified in milk carbohydrate of the red kangaroo. The presence of fucosyl oligosaccharides is a significant feature of koala milk, in which it differs from milk of the tammar wallaby and the red kangaroo.

  2. Asparagine-linked oligosaccharides on lutropin, follitropin, and thyrotropin: distributions of sulfated and sialylated oligosaccharides on bovine, ovine, and human pituitary glycoprotein hormones

    SciTech Connect

    Green, E.D.; Baenziger, J.U.

    1988-01-05

    The asparagine-linked oligosaccharides on the pituitary glycoprotein hormones lutropin (LH), follitropin (FSH), and thyrotropin (TSH) consist of a heterogeneous array of neutral, sulfated, sialylated, and sulfated/sialylated structures. In this study, the authors determined the relative quantities of the various asparagine-linked oligosaccharides on LH, FSH, and TSH from these three animal species. The proportions of sulfated versus sialylated oligosaccharides varied markedly among the different hormones. Both hormone- and animal species-specific differences in the types and distributions of sulfated, sialylated, and sulfated/sialylated structures were evident. In particular, LH and FSH, which are synthesized in the same pituitary cell and bear ..cap alpha..-subunits with the identical amino acid sequence, contained significantly different distributions of sulfated and sialylated oligosaccharides. For all three animal species, the ratio of sialylated to sulfated oligosaccharides differed by >10-fold for LH and FSH, with sulfated structures dominating on LH and sialylated structures on FSH. Sialylated oligosaccharides were also heterogeneous with respect to sialic acid linkage (..cap alpha..2,3 versus ..cap alpha..2,6). The differences in oligosaccharide structures among the various pituitary glycoprotein hormones as well as among the various glycosylation sites within a single hormone support the hypothesis that glycosylation may serve important functional roles in the expression and/or regulation of hormone bioactivity.

  3. Complex carbohydrates of red wine: characterization of the extreme diversity of neutral oligosaccharides by ESI-MS.

    PubMed

    Doco, Thierry; Williams, Pascale; Meudec, Emmanuelle; Cheynier, Véronique; Sommerer, Nicolas

    2015-01-21

    The major neutral oligosaccharides of a Carignan red wine have been characterized for the first time. The oligosaccharides were prepared after removal of phenolic compounds by polyamide chromatography and of polysaccharides by alcohol precipitation and then were fractionated by anion exchange and size-exclusion chromatography. In a second step, the glycosyl composition and linkages of wine oligosaccharides were determined. Oligosaccharide fractions were analyzed by mass spectrometry (MS) with an electrospray ionization (ESI) source and an ion trap mass analyzer after separation by hydrophilic interaction liquid chromatography on a Nucleodur HILIC column (zwitterionic sulfoalkyl betaine stationary phase). Glycosyl residue composition analysis showed the predominant presence of arabinose, with galactose, rhamnose, and mannose in lower proportion. Neutral oligosaccharides were present at a concentration of 185 mg/L in this wine. The MS spectra in the negative ion mode of the oligosaccharide fractions showed a series of oligosaccharidic structures corresponding to oligo-arabinans often linked to the basic unit α-l-Rhap-(1 → 4)-α-d-GalpA. The wine oligosaccharides identified correspond to arabino-oligosaccharides, rhamno-arabino-oligosaccharides, and different rhamnogalacturonan-arabino-oligosaccharides with DP ranging from 5 to 49, resulting from the degradation of grape cell wall pectins. Oligosaccharides have an extreme diversity, with more than 100 peaks detected in HPLC-ESI-MS spectra corresponding each to at least one oligosaccharidic structure.

  4. In vitro fermentation profiles, gas production rates, and microbiota modulation as affected by certain fructans, galactooligosaccharides, and polydextrose.

    PubMed

    Hernot, David C; Boileau, Thomas W; Bauer, Laura L; Middelbos, Ingmar S; Murphy, Michael R; Swanson, Kelly S; Fahey, George C

    2009-02-25

    It is of interest to benefit from the positive intestinal health outcomes of prebiotic consumption but with minimal gas production. This study examined gas production potential, fermentation profile, and microbial modulation properties of several types of oligosaccharides. Substrates studied included short-chain, medium-chain, and long-chain fructooligosaccharides, oligofructose-enriched inulin, galactooligosaccharide, and polydextrose. Each substrate was fermented in vitro using human fecal inoculum, and fermentation characteristics were quantified at 0, 4, 8, and 12 h. Gas and short-chain fatty acid (SCFA) production data showed that short-chain oligosaccharides were more rapidly fermented and produced more SCFA and gas than substrates with greater degrees of polymerization. Lactobacilli increased similarly among substrates. Short-chain oligosaccharides fermentation resulted in the greatest increase in bifidobacteria concentrations. Mixing short- and long-chain oligosaccharides attenuated short-chain oligosaccharide fermentation rate and extent. This study provides new information on the fermentation characteristics of some oligosaccharides used in human nutrition.

  5. Characterization of the oligosaccharides assembled on the Pichia pastoris-expressed recombinant aspartic protease.

    PubMed

    Montesino, R; Nimtz, M; Quintero, O; García, R; Falcón, V; Cremata, J A

    1999-10-01

    Aspartic protease, widely used as a milk-coagulating agent in industrial cheese production, contains three potential N-glycosylation sites. In this study, we report the characterization of N-linked oligosaccharides on recombinant aspartic protease secreted from the methylotrophic yeast Pichia pastoris using a combination of mass spectrometric, 2D chromatographic, chemical and enzymatic methods. The carbohydrates from site I (Asn79) were found to range from Man6-17GlcNAc2 with 50% bearing a phospho-diester-motif, site II (Asn113) was not occupied and site III (Asn188) contained mostly uncharged species ranging from Man-13GlcNAc2. These charged groups are not affecting the transport through the secretion pathway of the recombinant glycoprotein. Changes from a molasses-based medium to a minimal salts-based medium led to a clear reduction of the degree of phosphorylation of the N-glycan population.

  6. Characterization of sugar beet pectic-derived oligosaccharides obtained by enzymatic hydrolysis.

    PubMed

    Combo, Agnan Marie Michel; Aguedo, Mario; Quiévy, Nicolas; Danthine, Sabine; Goffin, Dorothée; Jacquet, Nicolas; Blecker, Christophe; Devaux, Jacques; Paquot, Michel

    2013-01-01

    Three pectic oligosaccharides (POS) obtained by enzymatic hydrolysis of sugar beet pectin by combining endopolygalacturonase and pectinmethylesterase, were characterized using high performance liquid chromatography, thermogravimetric analysis, Fourier transform infrared spectroscopy, differential scanning calorimetry and X-ray diffraction. According to chromatographic analyses, POS are composed of mixture of polymers with different molecular weights and different galacturonic acid contents. The thermal analysis showed no major variation in thermal behavior regarding POS composition but showed that POS were more sensitive to thermal degradation than the parent pectin as well as the deesterified pectin. No change in composition of the gaseous products was obtained through TGA-FTIR analysis. The X-ray pattern of POS clearly indicated a considerable decrease in crystallinity when compared to the native pectin. Thus, thermal characterization of POS may have practical repercussions if the formulation in which POS is incorporated is submitted to a high temperature treatment. Copyright © 2012 Elsevier B.V. All rights reserved.

  7. Characteristics of Lipo-Oligosaccharide Loci of Campylobacter jejuni Isolates Associated with Guillain-Barré Syndrome from Hebei, China

    PubMed Central

    Jiang, Hai; Zhang, Mao-Jun; Liu, Rui-Chun; Tian, Xin-Ying; Gu, Yi-Xin; Zhang, Jian-Zhong

    2010-01-01

    Ganglioside mimicry by C.jejuni lipo-oligosaccharides (LOS) could induce the production of autoantibodies against gangliosides and the development of Guillain-Barré syndrome (GBS). The LOS biosynthesis region exhibits significant variation with different strains. Using PCR amplifications of genes from published LOS loci and sequencing the LOS biosynthesis loci, the eight GBS-associated C. jejuni strains from HeBei could be classified into four classes. The expression of sialylated LOS structures (class A) or non-sialylated LOS structures(class F, H and P) in the C. jejuni LOS is considered to be two different factors for the induction of GBS. PMID:20480006

  8. Thermotoga maritima maltosyltransferase, a novel type of maltodextrin glycosyltransferase acting on starch and malto-oligosaccharides.

    PubMed

    Meissner, H; Liebl, W

    1998-12-15

    A novel enzyme acting on starch and malto-oligosaccharides was identified and characterised. The non-hydrolytic enzyme, designated maltosyltransferase (MTase), of the hyperthermophilic bacterium Thermotoga maritima MSB8 disproportionates malto-oligosaccharides via glycosyl transfer reactions. The enzyme has a unique transfer specificity strictly confined to the transfer of maltosyl units. Incubation of MTase with starch or its constituents. i.e. amylose and amylopectin, led to the formation of a set of multiples of maltose (i.e. maltose, maltotetraose, maltohexaose etc.). Malto-oligosaccharides with a degree of polymerization (DP) X were disproportionated to products with a DP of X +/- 2n (with X > or = 3 and n = 0,1,2,...). Maximum activity in a 10-min assay was recorded at pH 6.5 and 85-90 degrees C. The enzyme displayed extraordinary resistance to thermal inactivation. For example, at 90, 85, and 70 degrees C (pH 6.5, 0.34 mg ml-1 protein), MTase half-lives of about 2.5 h, 17 h, and 21 days, respectively, were recorded. The gene for MTase, designated mmtA, was isolated from a gene library of T. maritima strain MSB8. Analysis of the MTase primary structure as deduced from the nucleotide sequence of mmtA revealed that the enzyme is not closely related to known protein sequences. However, low-level local similarity between MTase and the alpha-amylase enzyme family (glycosyl hydrolase family 13) was detected, including conserved acidic residues essential for catalysis. Therefore, MTase should be assigned to this family. Based on detailed sequence analyses and comparison with amylolytic enzymes of known crystal structure we propose that MTase contains a (beta/alpha)8-fold as the core supersecondary structure which is typical for the alpha-amylase family. On the other hand, MTase is unique in that it lacks several residues highly conserved throughout this family. Also, MTase possesses an extraordinarily large domain B (a domain typical for the alpha-amylase family

  9. Protection of probiotic bacteria in synbiotic matrices

    USDA-ARS?s Scientific Manuscript database

    Probiotics, like Lactobacillus acidophilus, Lactobacillus reuteri, Bifidobacterium breve, Bifidobacterium longum, when encapsulated with prebiotic fibers such as fructo-oligosaccharides (FOS), inulin (I) and pectic-oligosaccharides (POS), formed a synbiotic matrix system that protected the bacteria ...

  10. Detection of milk oligosaccharides in plasma of infants

    PubMed Central

    Ruhaak, L. Renee; Stroble, Carol; Underwood, Mark A.; Lebrilla, Carlito B.

    2014-01-01

    Human Milk Oligosaccharides (HMO) are one of the major components of human milk. HMO are non-digestible by the human gut, where they are known to play important functions as prebiotics and decoys for binding pathogens. Moreover, it has been proposed that HMO may provide sialic acids to the infant that are important in brain development, however this would require absorption of HMO into the bloodstream. HMO have consistently been found in the urine of humans and other mammals, suggesting systemic absorption. Here we present a procedure for the profiling of milk oligosaccharides (MO) in plasma samples obtained from 13 term infants hospitalized for surgery for congenital heart disease. The method comprises protein denaturation, oligosaccharide reduction and porous graphitized carbon solid phase extraction for purification followed by analysis using nHPLC-PGC-chip-TOF-MS. Approximately 15 free MO were typically observed in the plasma of human infants, including LNT, LDFP, LNFT, 3’SL, 6’SL, 3’SLN and 6’SLN, of which the presence was confirmed using fragmentation studies. A novel third isomer of SLN, not found in human or bovine milk was also consistently detected. Differences in the free MO profiles were observed between infants that were totally formula-fed and infants that received at least some part breast milk. Our results indicate that free MO similar in structure to those found in human milk and urine are present in the blood of infants. The method and results presented here will facilitate further research toward the possible roles of free MO in the development of the infant. PMID:25059723

  11. Understanding the fundamental mechanism behind accumulation of oligosaccharides during high solids loading enzymatic hydrolysis

    USDA-ARS?s Scientific Manuscript database

    During enzymatic hydrolysis of biomass, polysaccharides are cleaved by glycosyl hydrolases to soluble oligosaccharides and further hydrolyzed by ß-glucosidase, ß-xylosidase and other enzymes to monomeric sugars. However, commercial enzyme mixtures do not hydrolyze all of these oligosaccharides and v...

  12. Predicted configurations of oligosaccharide extensions in the lipooligosaccharide of nontypeable Haemophilus influenzae isolates.

    PubMed

    McCrea, Kirk W; Xie, Jingping; Daniel, Deborah; Ulrich-Lewis, Justin Theophilus; Zhang, Lixin

    2014-07-01

    Lipooligosaccharide configurations were predicted in nontypeable Haemophilus influenzae isolates based on the presence of seven oligosaccharide extension-initiating genes (or alleles). Predicted configurations with 2 to 3 oligosaccharide extensions were more prevalent among middle ear than throat strains. In addition, strains with these configurations averaged higher levels of serum resistance than strains with other configurations.

  13. Feruloyl oligosaccharides from cell walls of suspension-cultured spinach cells and sugar beet pulp.

    PubMed

    Ishii, T

    1994-06-01

    Cell walls of suspension-cultured spinach cells and sugar beet pulp were separately hydrolyzed with Driselase. A feruloyl arabinobiose was isolated from both spinach cells and sugar beet. Four feruloyl oligosaccharides were obtained from sugar beet. The four oligosaccharides were characterized by NMR spectroscopy, methylation analysis and FAB-MS.

  14. Structural Basis for Norovirus Inhibition by Human Milk Oligosaccharides

    PubMed Central

    Weichert, Stefan; Koromyslova, Anna; Singh, Bishal K.; Hansman, Satoko; Jennewein, Stefan; Schroten, Horst

    2016-01-01

    Histo-blood group antigens (HBGAs) are important binding factors for norovirus infections. We show that two human milk oligosaccharides, 2′-fucosyllactose (2′FL) and 3-fucosyllactose (3FL), could block norovirus from binding to surrogate HBGA samples. We found that 2′FL and 3FL bound at the equivalent HBGA pockets on the norovirus capsid using X-ray crystallography. Our data revealed that 2′FL and 3FL structurally mimic HBGAs. These results suggest that 2′FL and 3FL might act as naturally occurring decoys in humans. PMID:26889023

  15. Hyaluronidase and Hyaluronan Oligosaccharides Promote Neurological Recovery after Intraventricular Hemorrhage

    PubMed Central

    Vinukonda, Govindaiah; Dohare, Preeti; Arshad, Arslan; Zia, Muhammad T.; Panda, Sanjeet; Korumilli, Ritesh; Kayton, Robert; Hascall, Vincent C.; Lauer, Mark E.

    2016-01-01

    Intraventricular hemorrhage (IVH) in premature infants results in inflammation, arrested oligodendrocyte progenitor cell (OPC) maturation, and reduced myelination of the white matter. Hyaluronan (HA) inhibits OPC maturation and complexes with the heavy chain (HC) of glycoprotein inter-α-inhibitor to form pathological HA (HC–HA complex), which exacerbates inflammation. Therefore, we hypothesized that IVH would result in accumulation of HA, and that either degradation of HA by hyaluronidase treatment or elimination of HCs from pathological HA by HA oligosaccharide administration would restore OPC maturation, myelination, and neurological function in survivors with IVH. To test these hypotheses, we used the preterm rabbit model of glycerol-induced IVH and analyzed autopsy samples from premature infants. We found that total HA levels were comparable in both preterm rabbit pups and human infants with and without IVH, but HA receptors—CD44, TLR2, TLR4—were elevated in the forebrain of both humans and rabbits with IVH. Hyaluronidase treatment of rabbits with IVH reduced CD44 and TLR4 expression, proinflammatory cytokine levels, and microglia infiltration. It also promoted OPC maturation, myelination, and neurological recovery. HC–HA and tumor necrosis factor-stimulated gene-6 were elevated in newborns with IVH; and depletion of HC–HA levels by HA oligosaccharide treatment reduced inflammation and enhanced myelination and neurological recovery in rabbits with IVH. Hence, hyaluronidase or HA oligosaccharide treatment represses inflammation, promotes OPC maturation, and restores myelination and neurological function in rabbits with IVH. These therapeutic strategies might improve the neurological outcome of premature infants with IVH. SIGNIFICANCE STATEMENT Approximately 12,000 premature infants develop IVH every year in the United States, and a large number of survivors with IVH develop cerebral palsy and cognitive deficits. The onset of IVH induces inflammation

  16. Conformational analysis of oligosaccharides and polysaccharides using molecular dynamics simulations.

    PubMed

    Frank, Martin

    2015-01-01

    Complex carbohydrates usually have a large number of rotatable bonds and consequently a large number of theoretically possible conformations can be generated (combinatorial explosion). The application of systematic search methods for conformational analysis of carbohydrates is therefore limited to disaccharides and trisaccharides in a routine analysis. An alternative approach is to use Monte-Carlo methods or (high-temperature) molecular dynamics (MD) simulations to explore the conformational space of complex carbohydrates. This chapter describes how to use MD simulation data to perform a conformational analysis (conformational maps, hydrogen bonds) of oligosaccharides and how to build realistic 3D structures of large polysaccharides using Conformational Analysis Tools (CAT).

  17. Quantitation and structures of oligosaccharide chains in human trachea mucin glycoproteins.

    PubMed

    Sangadala, S; Bhat, U R; Mendicino, J

    1992-12-02

    Human respiratory mucin glycoproteins from patients with cystic fibrosis were purified and oligosaccharide chains were released by treatment with alkaline borohydride. A neutral oligosaccharide alditol fraction was isolated from mucin obtained from a patient with A blood group determinant by chromatography on DEAE-cellulose and individual oligosaccharide chains were then isolated by gel filtration on BioGel P-6 columns and high performance liquid chromatography with gradient and isocratic solvent systems. The structures of the purified oligosaccharides were determined by methylation analysis, sequential glycosidase digestion and 'H-NMR spectroscopy. The amount of each chain was determined by compositional analysis. A wide array of discrete branched oligosaccharide structures that contain from 3 to 22 sugar residues were found. Many of the oligosaccharides are related and appear to be precursors of larger chains. The predominant branched oligosaccharides which accumulate contain terminal blood group H (Fuc alpha 2Ga1 beta 4) or blood group A (Fuc alpha 2(Ga1NAc alpha 3) (Ga1 beta 4) determinants which stop further branching and chain elongation. The elongation of oligosaccharide chains in respiratory mucins occurs on the beta 3-linked G1cNAc at branch points, whereas the beta 6-linked G1cNAc residue ultimately forms short side chains with a Fuc alpha 2(Ga1NAc alpha 3) Ga1 beta 4 G1cNAc beta 6 structure in individuals with A blood group determinant. The results obtained in the current studies further suggest that even higher molecular weight oligosaccharide chains with analogous branched structures are present in some human respiratory mucin glycoproteins. Increasing numbers of the repeating sequence shown in the oligosaccharide below is present in the higher molecular weight chains. [formula: see text] This data in conjunction with our earlier observations on the extensive branching of these oligosaccharide chains helps to define and explain the enormous range of

  18. Structural basis of chitin oligosaccharide deacetylation.

    PubMed

    Andrés, Eduardo; Albesa-Jové, David; Biarnés, Xevi; Moerschbacher, Bruno M; Guerin, Marcelo E; Planas, Antoni

    2014-07-01

    Cell signaling and other biological activities of chitooligosaccharides (COSs) seem to be dependent not only on the degree of polymerization, but markedly on the specific de-N-acetylation pattern. Chitin de-N-acetylases (CDAs) catalyze the hydrolysis of the acetamido group in GlcNAc residues of chitin, chitosan, and COS. A major challenge is to understand how CDAs specifically define the distribution of GlcNAc and GlcNH2 moieties in the oligomeric chain. We report the crystal structure of the Vibrio cholerae CDA in four relevant states of its catalytic cycle. The two enzyme complexes with chitobiose and chitotriose represent the first 3D structures of a CDA with its natural substrates in a productive mode for catalysis, thereby unraveling an induced-fit mechanism with a significant conformational change of a loop closing the active site. We propose that the deacetylation pattern exhibited by different CDAs is governed by critical loops that shape and differentially block accessible subsites in the binding cleft of CE4 enzymes.

  19. Disruption and eradication of P. aeruginosa biofilms using nitric oxide-releasing chitosan oligosaccharides

    PubMed Central

    Reighard, Katelyn P.; Hill, David B.; Dixon, Graham A.; Worley, Brittany; Schoenfisch, Mark H.

    2015-01-01

    Biofilm disruption and eradication were investigated as a function of nitric oxide- (NO) releasing chitosan oligosaccharide dose with results compared to control (ie non-NO-releasing) chitosan oligosaccharides and tobramycin. Quantification of biofilm expansion/contraction and multiple-particle tracking microrheology were used to assess the structural integrity of the biofilm before and after antibacterial treatment. While tobramycin had no effect on the physical properties of the biofilm, NO-releasing chitosan oligosaccharides exhibited dose-dependent behavior with biofilm degradation. Control chitosan oligosaccharides increased biofilm elasticity, indicating that the scaffold may mitigate the biofilm disrupting power of nitric oxide somewhat. The results from this study indicate that nitric oxide-releasing chitosan oligosaccharides act as dual-action therapeutics capable of eradicating and physically disrupting P. aeruginosa biofilms. PMID:26610146

  20. Preparation and characterization of 15N-enriched, size-defined heparan sulfate precursor oligosaccharides

    PubMed Central

    Sigulinsky, Crystal; Babu, Ponnusamy; Victor, Xylophone V.; Kuberan, Balagurunathan

    2009-01-01

    We report the preparation of size-defined [15N]N-acetylheparosan oligosaccharides from Escherichia coli-derived 15N-enriched N-acetylheparosan. Optimized growth conditions of E. coli in minimal media containing 15NH4Cl yielded [15N]N-acetylheparosan on a preparative scale. Depolymerization of [15N]N-acetylheparosan by heparitinase I yielded resolvable, even-numbered oligosaccharides ranging from disaccharide to icosaccharide. Anion-exchange chromatography-assisted fractionation afforded size-defined [15N]N-acetylheparosan oligosaccharides identifiable by ESI-TOFMS. These isotopically labeled oligosaccharides will prove to be valuable research tools for the chemoenzymatic synthesis of heparin and heparan sulfate oligosaccharides and for the study of their structural biology. PMID:19945695

  1. The chitin catabolic cascade in the marine bacterium Vibrio cholerae: characterization of a unique chitin oligosaccharide deacetylase.

    PubMed

    Li, Xibing; Wang, Lai-Xi; Wang, Xuesong; Roseman, Saul

    2007-12-01

    Chitin, one of the most abundant organic substances in nature, is consumed by marine bacteria, such as Vibrio cholerae, via a multitude of tightly regulated genes (Li and Roseman 2004, Proc Natl Acad Sci USA. 101:627-631). One such gene, cod, is reported here. It encodes a chitin oligosaccharide deacetylase (COD), when cells are induced by chitobiose, (GlcNH(2))(2), or crude crab shells. COD was molecularly cloned (COD-6His), overproduced, and purified to apparent homogeneity. COD is secreted at all stages of growth by induced V. cholerae. The gene sequence predicts a 26 N-terminal amino acid signal peptide not found in the isolated protein. COD is very active with chitin oligosaccharides, is virtually inactive with GlcNAc, and slightly active with colloidal ([(3)H]-N-acetyl)-chitin. The oligosaccharides are converted almost quantitatively to products lacking one acetyl group. The latter were characterized by mass spectrometry (ESI-MS), and treatment with nitrous acid. COD catalyzes the following reactions (n = 2-6): (GlcNAc)(n)--> GlcNAc-GlcNH(2)-(GlcNAc)(n-2) + Ac(-). That is, COD hydrolyzes the N-acetyl groups attached to the penultimate GlcNAc residue. The gene bank sequence data show that cod is highly conserved in Vibrios and Photobacteria. One such gene encodes a deacetylase isolated from V. alginolytics (Ohishi et al. 1997, Biosci Biotech Biochem. 61:1113-1117; Ohishi et al. 2000, J Biosci Bioeng. 90:561-563), that is specific for (GlcNAc)(2), but inactive with higher oligosaccharides. The COD enzymatic products, GlcNAc-GlcNH(2)-(GlcNAc)(n), closely resemble those obtained by hydrolysis of the chitooligosaccharides with Nod B: GlcNH(2)-(GlcNAc)(3-4). The latter are key intermediates in the biosynthesis of Nod factors, critically important in communications between the symbiotic nitrogen fixing bacteria and plants. Conceivably, the COD products play equally important roles in cellular communications that remain to be defined.

  2. Evaluation of fermentable oligosaccharides in diets fed to dogs in comparison to fiber standards.

    PubMed

    Middelbos, I S; Fastinger, N D; Fahey, G C

    2007-11-01

    Blends of fermentable oligosaccharides in combination with nonfermentable fiber, cellulose, were evaluated for their ability to serve as dietary fibers in dog foods. Using a 6 x 6 Latin square design, 6 diets were evaluated that contained either no supplemental fiber, beet pulp, cellulose, or blends of cellulose, fructooligosaccharides, and yeast cell wall added at 2.5% of the diet. Six ileal-cannulated dogs were fed 175 g of their assigned diet twice daily. Chromic oxide served as a digestibility marker. Nutrient digestibility, fecal microbial populations, fermentative end products, and immunological indices were measured. Total tract DM and OM digestibilities were lowest (P < 0.05) for the cellulose treatment. Crude protein digestibility was lower (P < 0.05) for the treatments containing carbohydrate blends. The cellulose treatment had the lowest (P < 0.05) concentration of bacteria, and all diets containing fermentable fiber had greater (P < 0.05) fecal bifidobacteria concentrations compared with the diets without supplemental fermentable fiber. Lactobacilli concentrations tended to be greater (P < 0.08) in treatments containing fermentable fiber compared with the cellulose treatment. Bifidobacteria and lactobacilli concentrations were similar for the beet pulp treatment compared with the fermentable oligosaccharide blends. Total fecal short-chain fatty acid concentration was greater for the beet pulp treatment (P < 0.05) compared with the control and cellulose treatments. The treatments containing fermentable fiber had greater (P < 0.05) fecal butyrate concentrations compared with cellulose and control treatments. Immune indices were not affected by treatment. Our results suggest that dog foods containing blends of fermentable and nonfermentable carbohydrates produce similar physiological results as dog food containing beet pulp as a fiber source. Therefore, blends of these carbohydrates could be useful substitutes for beet pulp in dog foods.

  3. Attenuation of food allergy symptoms following treatment with human milk oligosaccharides in a mouse model.

    PubMed

    Castillo-Courtade, L; Han, S; Lee, S; Mian, F M; Buck, R; Forsythe, P

    2015-09-01

    The prebiotic nature of human milk oligosaccharides (HMOs) and increasing evidence of direct immunomodulatory effects of these sugars suggest that they may have some therapeutic potential in allergy. Here, we assess the effect of two HMOs, 2'-fucosyllactose and 6'-sialyllactose, on symptomatology and immune responses in an ovalbumin-sensitized mouse model of food allergy. The effects of oral treatment with 2'-fucosyllactose and 6'-sialyllactose on anaphylactic symptoms induced by oral ovalbumin (OVA) challenge in sensitized mice were investigated. Mast cell functions in response to oral HMO treatment were also measured in the passive cutaneous anaphylaxis model, and direct effects on IgE-mediated degranulation of mast cells were assessed. Daily oral treatment with 2'-fucosyllactose or 6'-sialyllactose attenuated food allergy symptoms including diarrhea and hypothermia. Treatment with HMOs also suppressed antigen-induced increases in mouse mast cell protease-1 in serum and mast cell numbers in the intestine. These effects were associated with increases in the CD4(+) CD25(+) IL-10(+) cell populations in the Peyer's patches and mesenteric lymph nodes, while 6'-sialyllactose also induced increased IL-10 and decreased TNF production in antigen-stimulated splenocytes. Both 2'-fucosyllactose and 6'-sialyllactose reduced the passive cutaneous anaphylaxis response, but only 6'-sialyllactose directly inhibited mast cell degranulation in vitro, at high concentrations. Our results suggest that 2'-fucosyllactose and 6'-sialyllactose reduce the symptoms of food allergy through induction of IL-10(+) T regulatory cells and indirect stabilization of mast cells. Thus, human milk oligosaccharides may have therapeutic potential in allergic disease. © 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  4. Carbohydrate-deficient glycoprotein syndrome: not an N-linked oligosaccharide processing defect, but an abnormality in lipid-linked oligosaccharide biosynthesis?

    PubMed Central

    Powell, L D; Paneerselvam, K; Vij, R; Diaz, S; Manzi, A; Buist, N; Freeze, H; Varki, A

    1994-01-01

    The carbohydrate-deficient glycoprotein syndrome (CDGS) is a developmental disease associated with an abnormally high isoelectric point of serum transferrin. Carbohydrate analyses of this glycoprotein initially suggested a defect in N-linked oligosaccharide processing, although more recent studies indicate a defect in the attachment of these sugar chains to the protein. We studied both serum glycoproteins and fibroblast-derived [2-3H]mannose-labeled oligosaccharides from CDGS patients and normal controls. While there was a decrease in the glycosylation of serum glycoproteins of affected individuals, differences were not seen in either monosaccharide composition or oligosaccharide structures. The lectin-binding profiles of glycopeptides from [2-3H]-mannose-labeled fibroblasts were likewise indistinguishable. However, the incorporation of [2-3H]mannose into both glycoproteins and the dolichol-linked oligosaccharide precursor was significantly reduced. Thus, at least in some patients, CDGS is not due to a defect in processing of N-linked oligosaccharides, but rather to defective synthesis and transfer of nascent dolichol-linked oligosaccharide precursors. This abnormality could result in both a failure to glycosylate some sites on some proteins, as well as secondary abnormalities in overall glycoprotein processing and/or function. PMID:7962535

  5. Asparagine-linked oligosaccharides on lutropin, follitropin, and thyrotropin: structural elucidation of the sulfated and sialylated oligosaccharides on bovine, ovine, and human pituitary glycoprotein hormones

    SciTech Connect

    Green, E.D.; Baenziger, J.U.

    1988-01-05

    The authors have elucidated the structures of the anionic asparagine-linked oligosaccharides present on the glycoprotein hormones lutropin (luteinizing hormone), follitropin (follicle-stimulating hormone), and thyrotropin (thyroid-stimulating hormone). Purified hormones, isolated from bovine, ovine, and human pituitaries, were digested with N-glycanase, and the released oligosaccharides were reduced with NaB(/sup 3/H)/sub 4/. The /sup 3/H-labeled oligosaccharides from each hormone were then fractionated by anion-exchange high performance liquid chromatography (HPLC) into populations differing in the number of sulfate and/or sialic acid moieties. The sulfated, sialylated, and sulfated/sialylated structures, which together comprised 67-90% of the asparagine-linked oligosaccharides on the pituitary glycoprotein hormones, were highly heterogeneous and displayed hormone- as well as animal species-specific features. A previously uncharacterized dibranched oligosaccharide, bearing one residue each of sulfate and sialic acid, was found on all of the hormones except bovine lutropin. In this study, they describe the purification and detailed structural characterizations of the sulfated, sialylated, and sulfated/sialylated oligosaccharides found on lutropin, follitropin, and thyrotropin from several animal species.

  6. Selective fermentation of gentiobiose-derived oligosaccharides by human gut bacteria and influence of molecular weight.

    PubMed

    Sanz, María Luz; Côté, Gregory L; Gibson, Glenn R; Rastall, Robert A

    2006-06-01

    Gentiooligosaccharides and alternansucrase gentiobiose acceptor products were fractionated by their degree of polymerization (DP) on a Bio-Gel P2 column. Fractions were characterized by matrix-assisted laser desorption ionization time-of-flight mass spectroscopy, and incubated with human faecal bacteria under anaerobic conditions at 37 degrees C. The growth of predominant gut bacteria on the oligosaccharides was evaluated by fluorescence in situ hybridization and a prebiotic index (PI) was calculated. Lower DP gentiooligosaccharides (DP2-3) showed the highest selectivity (PI of 4.89 and 3.40, respectively), whereas DP4-5 alternansucrase gentiobiose acceptor products generated the greatest values (PI of 5.87). The production of short-chain fatty acids was also determined during the time course of the reactions. The mixture of DP6-10 alternansucrase gentiobiose acceptor products generated the highest levels of butyric acid but the lowest levels of lactic acid. Generally, for similar molecular weights, alternansucrase gentiobiose acceptor products gave higher PI values than gentiooligosaccharides.

  7. The CebE/MsiK Transporter is a Doorway to the Cello-oligosaccharide-mediated Induction of Streptomyces scabies Pathogenicity

    PubMed Central

    Jourdan, Samuel; Francis, Isolde Maria; Kim, Min Jung; Salazar, Joren Jeico C.; Planckaert, Sören; Frère, Jean-Marie; Matagne, André; Kerff, Frédéric; Devreese, Bart; Loria, Rosemary; Rigali, Sébastien

    2016-01-01

    Streptomyces scabies is an economically important plant pathogen well-known for damaging root and tuber crops by causing scab lesions. Thaxtomin A is the main causative agent responsible for the pathogenicity of S. scabies and cello-oligosaccharides are environmental triggers that induce the production of this phytotoxin. How cello-oligosaccharides are sensed or transported in order to induce the virulent behavior of S. scabies? Here we report that the cellobiose and cellotriose binding protein CebE, and MsiK, the ATPase providing energy for carbohydrates transport, are the protagonists of the cello-oligosaccharide mediated induction of thaxtomin production in S. scabies. Our work provides the first example where the transport and not the sensing of major constituents of the plant host is the central mechanism associated with virulence of the pathogen. Our results allow to draw a complete pathway from signal transport to phytotoxin production where each step of the cascade is controlled by CebR, the cellulose utilization regulator. We propose the high affinity of CebE to cellotriose as possible adaptation of S. scabies to colonize expanding plant tissue. Our work further highlights how genes associated with primary metabolism in nonpathogenic Streptomyces species have been recruited as basic elements of virulence in plant pathogenic species. PMID:27250236

  8. Will isomalto-oligosaccharides, a well-established functional food in Asia, break through the European and American market? The status of knowledge on these prebiotics.

    PubMed

    Goffin, Dorothee; Delzenne, Nathalie; Blecker, Christophe; Hanon, Emilien; Deroanne, Claude; Paquot, Michel

    2011-05-01

    This critical review article presents the current state of knowledge on isomalto-oligosaccharides, some well known functional oligosaccharides in Asia, to evaluate their potential as emergent prebiotics in the American and European functional food market. It includes first a unique inventory of the different families of compounds which have been considered as IMOs and their specific structure. A description has been given of the different production methods including the involved enzymes and their specific activities, the substrates, and the types of IMOs produced. Considering the structural complexity of IMO products, specific characterization methods are described, as well as purification methods which enable the body to get rid of digestible oligosaccharides. Finally, an extensive review of their techno-functional and nutritional properties enables placing IMOs inside the growing prebiotic market. This review is of particular interest considering that IMO commercialization in America and Europe is a topical subject due to the recent submission by Bioneutra Inc. (Canada) of a novel food file to the UK Food Standards Agency, as well as several patents for IMO production.

  9. Significance of galactinol and raffinose family oligosaccharide synthesis in plants

    PubMed Central

    Sengupta, Sonali; Mukherjee, Sritama; Basak, Papri; Majumder, Arun L.

    2015-01-01

    Abiotic stress induces differential expression of genes responsible for the synthesis of raffinose family of oligosaccharides (RFOs) in plants. RFOs are described as the most widespread D-galactose containing oligosaccharides in higher plants. Biosynthesis of RFOs begin with the activity of galactinol synthase (GolS; EC 2.4.1.123), a GT8 family glycosyltransferase that galactosylates myo-inositol to produce galactinol. Raffinose and the subsequent higher molecular weight RFOs (Stachyose, Verbascose, and Ajugose) are synthesized from sucrose by the subsequent addition of activated galactose moieties donated by Galactinol. Interestingly, GolS, the key enzyme of this pathway is functional only in the flowering plants. It is thus assumed that RFO synthesis is a specialized metabolic event in higher plants; although it is not known whether lower plant groups synthesize any galactinol or RFOs. In higher plants, several functional importance of RFOs have been reported, e.g., RFOs protect the embryo from maturation associated desiccation, are predominant transport carbohydrates in some plant families, act as signaling molecule following pathogen attack and wounding and accumulate in vegetative tissues in response to a range of abiotic stresses. However, the loss-of-function mutants reported so far fail to show any perturbation in those biological functions. The role of RFOs in biotic and abiotic stress is therefore still in debate and their specificity and related components remains to be demonstrated. The present review discusses the biology and stress-linked regulation of this less studied extension of inositol metabolic pathway. PMID:26379684

  10. Structural Determination and Daily Variations of Porcine Milk Oligosaccharides

    PubMed Central

    Tao, Nannan; Ochonicky, Karen L.; German, J. Bruce; Donovan, Sharon M.; Lebrilla, Carlito B.

    2010-01-01

    Free milk oligosaccharides (OS) is a major component of mammalian milk. Swine are important agricultural species and biomedical models. Despite their importance, little is known of the OS profile of porcine milk. Herein, the porcine milk glycome was elucidated and monitored over the entire lactation period by liquid chromatography profiling and structural determination with mass spectrometry. Milk was collected from second parity sows (n=3) at farrowing and on days 1, 4, 7 and 24 of lactation. Twenty-nine distinct porcine milk oligosaccharides (pMO) were identified. The pMO are highly sialylated, which is more similar to bovine milk than human milk OS. Six fucosylated pMO were detected at low levels in porcine milk, making it more similar to human milk than bovine. In general, the pMO content was highest in milk collected at farrowing and day 1 of lactation, decreased during early lactation, but then rose at day 24, however, the pMO displayed different patterns of variation across lactation. In summary, porcine milk contains both acidic (sialylated) and neutral (fucosylated) OS, but sialic-acid containing OS predominate throughout lactation. PMID:20369835

  11. Assessment of the two Helicobacter pylori alpha-1,3-fucosyltransferase ortholog genes for the large-scale synthesis of LewisX human milk oligosaccharides by metabolically engineered Escherichia coli.

    PubMed

    Dumon, Claire; Samain, Eric; Priem, Bernard

    2004-01-01

    We previously described a bacterial fermentation process for the in vivo conversion of lactose into fucosylated derivatives of lacto-N-neotetraose Gal(beta1-4)GlcNAc(beta1-3)Gal(beta1-4)Glc (LNnT). The major product obtained was lacto-N-neofucopentaose-V Gal(beta1-4)GlcNAc(beta1-3)Gal(beta1-4)[Fuc(alpha1-3)]Glc, carrying fucose on the glucosyl residue of LNnT. Only a small amount of oligosaccharides fucosylated on N-acetylglucosaminyl residues and thus carrying the LewisX group (Le(X)) was also produced. We report here a fermentation process for the large-scale production of Le(X) oligosaccharides. The two fucosyltransferase genes futA and futB of Helicobacter pylori (strain 26695) were compared in order to optimize fucosylation in vivo. futA was found to provide the best activity on the LNnT acceptor, whereas futB expressed a better Le(X) activity in vitro. Both genes were expressed to produce oligosaccharides in engineered Escherichia coli (E. coli) cells. The fucosylation pattern of the recombinant oligosaccharides was closely correlated with the specificity observed in vitro, FutB favoring the formation of Le(X) carrying oligosaccharides. Lacto-N-neodifucohexaose-II Gal(beta1-4)[Fuc(alpha1-3)]GlcNAc(beta1-3)Gal(beta1-4)[Fuc(alpha1-3)]Glc represented 70% of the total oligosaccharide amount of futA-on-driven fermentation and was produced at a concentration of 1.7 g/L. Fermentation driven by futB led to equal amounts of both lacto-N-neofucopentaose-V and lacto-N-neofucopentaose-II Gal(beta1-4)[Fuc(alpha1-3)]GlcNAc(beta1-3)Gal(beta1-4)Glc, produced at 280 and 260 mg/L, respectively. Unexpectedly, a noticeable proportion (0.5 g/L) of the human milk oligosaccharide 3-fucosyllactose Gal(beta1-4)[Fuc(alpha1-3)]Glc was produced in futA-on-driven fermentation, underlining the activity of fucosyltransferase FutA in E. coli and leading to a reassessment of its activity on lactose. All oligosaccharides produced by the products of both fut genes were natural compounds of

  12. Simultaneous synthesis of mixtures of lactulose and galacto-oligosaccharides and their selective fermentation.

    PubMed

    Guerrero, Cecilia; Vera, Carlos; Acevedo, Fernando; Illanes, Andrés

    2015-09-10

    Lactulose and galacto-oligosaccharides (GOS) are well recognized prebiotics derived from lactose. In the synthesis of lactulose with β-galactosidases GOS are also produced, but the ratio of lactulose and GOS in the product can be tuned at will, depending on the operation conditions, so to obtain an optimal product distribution in terms of prebiotic potential. The selectivity of fermentation of each carbohydrate alone as well as mixtures of both was determined using pH-controlled anaerobic batch cultures with faecal inoculum. Within the experimental range considered, lactulose/GOS molar ratio of 4 resulted in the highest selectivity for Bifidobacterium and Lactobacillus/Enterococcus, so this ratio was selected as the target for the synthesis of lactulose from fructose and lactose with Aspergillus oryzae β-galactosidase. Synthesis was optimized using response surface methodology, considering temperature, initial concentrations of acceptor sugars and fructose/lactose molar ratio as key variables, with the aim of maximizing lactulose yield at the optimal product distribution in terms of prebiotic potential (lactulose/GOS molar ratio of 4). Under optimal conditions (50°C, 50%w/w total initial concentrations of sugars and fructose/lactose molar ratio of 6.44), lactulose yield of 0.26g of lactulose produced per g of initial lactose was obtained at the optimal product distribution.

  13. Assignment of the Stereochemistry and Anomeric Configuration of Sugars within Oligosaccharides Via Overlapping Disaccharide Ladders Using MSn

    NASA Astrophysics Data System (ADS)

    Konda, Chiharu; Londry, Frank A.; Bendiak, Brad; Xia, Yu

    2014-08-01

    A systematic approach is described that can pinpoint the stereo-structures (sugar identity, anomeric configuration, and location) of individual sugar units within linear oligosaccharides. Using a highly modified mass spectrometer, dissociation of linear oligosaccharides in the gas phase was optimized along multiple-stage tandem dissociation pathways (MSn, n = 4 or 5). The instrument was a hybrid triple quadrupole/linear ion trap mass spectrometer capable of high-efficiency bidirectional ion transfer between quadrupole arrays. Different types of collision-induced dissociation (CID), either on-resonance ion trap or beam-type CID could be utilized at any given stage of dissociation, enabling either glycosidic bond cleavages or cross-ring cleavages to be maximized when wanted. The approach first involves optimizing the isolation of disaccharide units as an ordered set of overlapping substructures via glycosidic bond cleavages during early stages of MSn, with explicit intent to minimize cross-ring cleavages. Subsequently, cross-ring cleavages were optimized for individual disaccharides to yield key diagnostic product ions ( m/ z 221). Finally, fingerprint patterns that establish stereochemistry and anomeric configuration were obtained from the diagnostic ions via CID. Model linear oligosaccharides were derivatized at the reducing end, allowing overlapping ladders of disaccharides to be isolated from MSn. High confidence stereo-structural determination was achieved by matching MSn CID of the diagnostic ions to synthetic standards via a spectral matching algorithm. Using this MSn ( n = 4 or 5) approach, the stereo-structures, anomeric configurations, and locations of three individual sugar units within two pentasaccharides were successfully determined.

  14. Selective control of oligosaccharide transfer efficiency for the N-glycosylation sequon by a point mutation in oligosaccharyltransferase.

    PubMed

    Igura, Mayumi; Kohda, Daisuke

    2011-04-15

    Asn-linked glycosylation is the most ubiquitous posttranslational protein modification in eukaryotes and archaea, and in some eubacteria. Oligosaccharyltransferase (OST) catalyzes the transfer of preassembled oligosaccharides on lipid carriers onto asparagine residues in polypeptide chains. Inefficient oligosaccharide transfer results in glycoprotein heterogeneity, which is particularly bothersome in pharmaceutical glycoprotein production. Amino acid variation at the X position of the Asn-X-Ser/Thr sequon is known to modulate the glycosylation efficiency. The best amino acid at X is valine, for an archaeal Pyrococcus furiosus OST. We performed a systematic alanine mutagenesis study of the archaeal OST to identify the essential and dispensable amino acid residues in the three catalytic motifs. We then investigated the effects of the dispensable mutations on the amino acid preference in the N-glycosylation sequon. One residue position was found to selectively affect the amino acid preference at the X position. This residue is located within the recently identified DXXKXXX(M/I) motif, suggesting the involvement of this motif in N-glycosylation sequon recognition. In applications, mutations at this position may facilitate the design of OST variants adapted to particular N-glycosylation sites to reduce the heterogeneity of glycan occupancy. In fact, a mutation at this position led to 9-fold higher activity relative to the wild-type enzyme, toward a peptide containing arginine at X in place of valine. This mutational approach is potentially applicable to eukaryotic and eubacterial OSTs for the production of homogenous glycoproteins in engineered mammalian and Escherichia coli cells.

  15. Preparation and characterisation of the oligosaccharides derived from Chinese water chestnut polysaccharides.

    PubMed

    Wu, Sheng-Jun; Yu, Lin

    2015-08-15

    Hydrogen peroxide (H2O2) is a strong oxidant that cleaves glycosidic bonds in polysaccharides. In this study, the oligosaccharides were prepared by removing the starch from Chinese water chestnuts through hydrolysis using α-amylase and then hydrolysing the remaining polysaccharides with H2O2, during which the oligosaccharide yield was monitored. The yield of oligosaccharide was affected by reaction time, temperature, and H2O2 concentration. Extended reaction times, high temperatures, and high H2O2 concentrations decreased oligosaccharide yield. Under optimum conditions (i.e., reaction time of 4h, reaction temperature of 80°C, and 2.5% H2O2 concentration), the maximum oligosaccharide yield was 3.91%. The oligosaccharides derived from Chinese water chestnuts polysaccharides exhibited strong hydroxyl and 2,2-diphenyl-β-picrylhydrazyl radical scavenging activity when applied at a concentration of 100 μg/mL. The results indicate that the oligosaccharides derived from Chinese water chestnuts polysaccharides possessed good antioxidant properties and can be developed as a new dietary supplement and functional food.

  16. MALDI Q-TOF CID MS for Diagnostic Ion Screening of Human Milk Oligosaccharide Samples

    PubMed Central

    Jovanović, Marko; Tyldesley-Worster, Richard; Pohlentz, Gottfried; Peter-Katalinić, Jasna

    2014-01-01

    Human milk oligosaccharides (HMO) represent the bioactive components of human milk, influencing the infant’s gastrointestinal microflora and immune system. Structurally, they represent a highly complex class of analyte, where the main core oligosaccharide structures are built from galactose and N-acetylglucosamine, linked by 1–3 or 1–4 glycosidic linkages and potentially modified with fucose and sialic acid residues. The core structures can be linear or branched. Additional structural complexity in samples can be induced by endogenous exoglycosidase activity or chemical procedures during the sample preparation. Here, we show that using matrix-assisted laser desorption/ionization (MALDI) quadrupole-time-of-flight (Q-TOF) collision-induced dissociation (CID) as a fast screening method, diagnostic structural information about single oligosaccharide components present in a complex mixture can be obtained. According to sequencing data on 14 out of 22 parent ions detected in a single high molecular weight oligosaccharide chromatographic fraction, 20 different oligosaccharide structure types, corresponding to over 30 isomeric oligosaccharide structures and over 100 possible HMO isomers when biosynthetic linkage variations were taken into account, were postulated. For MS/MS data analysis, we used the de novo sequencing approach using diagnostic ion analysis on reduced oligosaccharides by following known biosynthetic rules. Using this approach, de novo characterization has been achieved also for the structures, which could not have been predicted. PMID:24743894

  17. Synthetic Oligosaccharide Stimulates and Stabilizes Angiogenesis: Structure–Function Relationships and Potential Mechanisms

    PubMed Central

    Mousa, S. A.; Feng, X.; Xie, J.; Du, Y.; Hua, Y.; He, H.; O’Connor, L.; Linhardt, R. J.

    2014-01-01

    To determine the proangiogenesis effect of series of saccharides and a synthetic oligosaccharide and potential mechanisms, an in vitro 3-dimensional endothelial cell sprouting (3D-ECS) assay and the chick chorioallantoic membrane (CAM) model were used. We demonstrated that a sulfated oligosaccharide significantly promotes the endothelial capillary network initiated by vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (b-FGF). Furthermore, although the capillary network initiated by VEGF and b-FGF lasts no more than 7 days, addition of a sulfated oligosaccharide significantly amplifies angiogenesis and stabilizes the capillary network of new blood vessels. In the CAM model, sulfated oligosaccharide also stimulated angiogenesis. In both the CAM and the 3D-ECS assay, structure–function studies reveal that increased saccharide chain length up to the hexa- to decasaccharide show optimal proangiogenesis efficacy. In addition, the sulfation and molecular shape (branched vs linear) of oligosaccharide are important for sustained proangiogenesis efficacy. Data indicate that chemically defined synthetic oligosaccharides can play an important role in regulation of capillary structure and stability, which may contribute to future advances in therapeutic angiogenesis. The proangiogenesis efficacy of an oligosaccharide is mediated via integrin αvβ3 and involves mitogen-activated protein kinase signaling mechanisms. PMID:16954815

  18. Comparative structural study of N-linked oligosaccharides of urinary and recombinant erythropoietins

    SciTech Connect

    Tsuda, E.; Goto, M.; Murakami, A.; Akia, K.; Ueda, M.; Kawanishi, G.; Takahashi, N.; Sasaki, R.; Chiba, H.; Ishihara, H.; Mori, M.

    1988-07-26

    The structures of the N-linked oligosaccharides of the urinary erythropoietin (u-EPOI) purified from urine of aplastic anemic patients were analyzed and compared with those for recombinant erythropoietin (r-EPO) prepared with baby hamster kidney (BHK) cells. Asparagine-linked neutral oligosaccharides were released from each EPO protein by N-oligosaccharide glycopeptidase (almond) digestion. The reducing ends of the oligosaccharide chains thus obtained were aminated with a fluorescent reagent, 2-aminopyridine, and the mixture of pyridylamino derivatives of the oligosaccharides was separated by high-performance liquid chromatography (HPLC) on an ODS silica column. More than 8 and 13 kinds of oligosaccharide fractions for u-EPO and r-EPO (BHK), respectively, were completely separated by the one-step HPLC procedure. The structure of each oligosaccharide thus isolated was analyzed by a combination of sequential exoglycosidase digestion and another kind of HPLC with an amide-silica column. Furthermore, high-resolution proton nuclear magnetic resonance (/sup 1/H NMR) spectroscopy and methylation analyses were carried out in the case of r-EPO (BHK).

  19. Comparative structural study of N-linked oligosaccharides of urinary and recombinant erythropoietins.

    PubMed

    Tsuda, E; Goto, M; Murakami, A; Akai, K; Ueda, M; Kawanishi, G; Takahashi, N; Sasaki, R; Chiba, H; Ishihara, H

    1988-07-26

    The structures of the N-linked oligosaccharides of the urinary erythropoietin (u-EPO) purified from urine of aplastic anemic patients were analyzed and compared with those for recombinant erythropoietin (r-EPO) prepared with baby hamster kidney (BHK) cells. Asparagine-linked neutral oligosaccharides were released from each EPO protein by N-oligosaccharide glycopeptidase (almond) digestion. The reducing ends of the oligosaccharide chains thus obtained were aminated with a fluorescent reagent, 2-aminopyridine, and the mixture of pyridylamino derivatives of the oligosaccharides was separated by high-performance liquid chromatography (HPLC) on an ODS silica column. More than 8 and 13 kinds of oligosaccharide fractions for u-EPO and r-EPO (BHK), respectively, were completely separated by the one-step HPLC procedure. The structure of each oligosaccharide thus isolated was analyzed by a combination of sequential exoglycosidase digestion and another kind of HPLC with an amide-silica column. Furthermore, high-resolution proton nuclear magnetic resonance (1H NMR) spectroscopy and methylation analyses were carried out in the case of r-EPO (BHK).(ABSTRACT TRUNCATED AT 250 WORDS)

  20. Effect of ensiling moist field bean (Vicia faba), pea (Pisum sativum) and lupine (Lupinus spp.) grains on the contents of alkaloids, oligosaccharides and tannins.

    PubMed

    Gefrom, A; Ott, E M; Hoedtke, S; Zeyner, A

    2013-12-01

    Ensiling legume grain may be an inexpensive and ecologically interesting method to produce a high-protein feed of local origin. The typically patchy maturation recommends harvesting and ensiling the seeds in moist condition. Developing a method for preserving legume grains harvested before maturation by lactic acid fermentation would have several advantages. Under laboratory conditions, crushed legume seeds of beans, peas and lupines with high moisture content of 35 % were ensiled with different additives (molasses and lactic acid bacteria). To characterize the final silages, contents of proximate nutrients and antinutritional factors (alkaloids, oligosaccharides, tannins) were analysed. The addition of lactic acid bacteria ensured a fast and pronounced lactic acid production and decreased contents of undesired fermentation products like ethanol. An additional use of molasses for ensilage did not provide a remarkable additional benefit. Excluding sugar and starch, the contents of proximate nutrients were not remarkably altered after ensiling. As an overall effect, lactic acid fermentation reduced tannins and oligosaccharides. It can be supposed that the oligosaccharides after breakdown of the complex molecules acted as a source of fermentable carbohydrates. A relevant reduction of alkaloids did not occur. The lactic acid fermentation of legume grains can be recommended as an appropriate method for conservation. With respect to the economic advantages and compared with methods of chemical preservation, the lactic acid fermentation of legume grains under anaerobic conditions is an environmentally compliant procedure and therefore also an option for organic farming.

  1. Kinetic analyses of retaining endo-(xylo)glucanases from plant and microbial sources using new chromogenic xylogluco-oligosaccharide aryl glycosides.

    PubMed

    Ibatullin, Farid M; Baumann, Martin J; Greffe, Lionel; Brumer, Harry

    2008-07-22

    A library of phenyl beta-glycosides of xylogluco-oligosaccharides was synthesized via a chemoenzymatic approach to produce new, specific substrates for xyloglucanases. Tamarind xyloglucan was completely hydrolyzed to four, variably galactosylated component oligosaccharides based on Glc 4 backbones, using a Trichoderma endo-glucanase mixture. Oligosaccharide complexity could be further reduced by beta-galactosidase treament. Subsequent per- O-acetylation, alpha-bromination, phase-transfer glycosylation, and Zemplen deprotection yielded phenyl glycosides of XXXG and XLLG oligosaccharides with a broad range of aglycon p K a values. Kinetic and product analysis of the action of the archetypal plant endo-xyloglucanase, Tropaeolum majus NXG1, on these compounds indicated that formation of the glycosyl-enzyme intermediate was rate-limiting in the case of phenol leaving groups with p K a values of >7, leading exclusively to substrate hydrolysis. Conversely, substrates with aglycon p K a values of 5.4 gave rise to a significant amount of transglycosylation products, indicating a change in the relative rates of formation and breakdown of the glycosyl-enzyme intermediate for these faster substrates. Notably, comparison of the initial rates of XXXG-Ar and XLLG-Ar conversion indicated that catalysis by TmNXG1 was essentially insensitive to the presence of galactose in the negative subsites for all leaving groups. More broadly, analysis of a selection of enzymes from CAZy families GH 5, 12, and 16 indicated that the phenyl glycosides are substrates for anomeric configuration-retaining endo-xyloglucanases but are not substrates for strict xyloglucan endo-transglycosylases (XETs). The relative activities of the GH 5, 12, and 16 endo-xyloglucanases toward GGGG-CNP, XXXG-CNP, and XLLG-CNP reflected those observed using analogous high molar mass polysaccharides. These new chromogenic substrates may thus find wide application in the discovery, screening, and detailed kinetic analysis

  2. Can an ancestral condition for milk oligosaccharides be determined? Evidence from the Tasmanian echidna (Tachyglossus aculeatus setosus).

    PubMed

    Oftedal, Olav T; Nicol, Stewart C; Davies, Noel W; Sekii, Nobuhiro; Taufik, Epi; Fukuda, Kenji; Saito, Tadao; Urashima, Tadasu

    2014-09-01

    The monotreme pattern of egg-incubation followed by extended lactation represents the ancestral mammalian reproductive condition, suggesting that monotreme milk may include saccharides of an ancestral type. Saccharides were characterized from milk of the Tasmanian echidna Tachyglossus aculeatus setosus. Oligosaccharides in pooled milk from late lactation were purified by gel filtration and high-performance liquid chromatography using a porous graphitized carbon column and characterized by (1)H NMR spectroscopy; oligosaccharides in smaller samples from early and mid-lactation were separated by ultra-performance liquid chromatography and characterized by negative electrospray ionization mass spectrometry (ESI-MS) and tandem collision mass spectroscopy (MS/MS) product ion patterns. Eight saccharides were identified by (1)H NMR: lactose, 2'-fucosyllactose, difucosyllactose (DFL), B-tetrasaccharide, B-pentasaccharide, lacto-N-fucopentaose III (LNFP3), 4-O-acetyl-3'-sialyllactose [Neu4,5Ac(α2-3)Gal(β1-4)Glc] and 4-O-acetyl-3'-sialyl-3-fucosyllactose [Neu4,5Ac(α2-3)Gal(β1-4)[Fuc(α1-3)]Glc]. Six of these (all except DFL and LNFP3) were present in early and mid-lactation per ESI-MS, although some at trace levels. Four additional oligosaccharides examined by ESI-MS and MS/MS are proposed to be 3'-sialyllactose [Neu5Ac(α2-3)Gal(β1-4)Glc], di-O-acetyl-3'-sialyllactose [Neu4,5,UAc3(α2-3)Gal(β1-4)Glc where U = 7, 8 or 9], 4-O-acetyl-3'-sialyllactose sulfate [Neu4,5Ac(α2-3)Gal(β1-4)GlcS, where position of the sulfate (S) is unknown] and an unidentified 800 Da oligosaccharide containing a 4-O-acetyl-3'-sialyllactose core. 4-O-acetyl-3'-sialyllactose was the predominant saccharide at all lactation stages. 4-O-Acetylation is known to protect sialyllactose from bacterial sialidases and may be critical to prevent microbial degradation on the mammary areolae and/or in the hatchling digestive tract so that sialyllactose can be available for enterocyte uptake. The ability to

  3. Human Milk Oligosaccharides: Evolution, Structures and Bioselectivity as Substrates for Intestinal Bacteria

    PubMed Central

    German, J. Bruce; Freeman, Samara L.; Lebrilla, Carlito B.; Mills, David A.

    2010-01-01

    Human milk contains a high concentration of diverse soluble oligosaccharides that are carbohydrate polymers formed from a relatively small number of different monosaccharides. Novel methods combining liquid chromatography with high resolution mass spectrometry have identified approximately 200 unique oligosaccharides structures varying from 3 to 22 sugars. The increasing structural complexity of oligosaccharides follows the general pattern of mammalian and primate evolution though the concentration and diversity of these structures in homo sapiens are strikingly more abundant. There is also considerable diversity among different human mothers in the structures of oligosaccharides. Milks from randomly selected mothers contain as few as 23 and as many as 130 different oligosaccharides. The functional implications of this diversity are not yet known. Despite the role of milk to serve as a sole nutrient source for mammalian infants, the majority of the oligosaccharides in milk are not digestible by human infants. This apparent paradox raises the obvious questions about the functions of these oligosaccharides and how their diverse molecular structures affect their functions. The nutritional function that is most frequently attributed to milk oligosaccharides is to serve as prebiotics –a form of indigestible carbohydrate that is selectively fermented by desirable gut microflora. This function was tested by purifying human milk oligosaccharides and providing these as the sole carbon source to various intestinal bacteria. Indeed, the selectively of providing the complex mixture of oligosaccharides pooled from dozens of human milk samples is remarkable. Among a variety of Bifidobacteria tested only Bifidobacteria longum biovar infantis was able to grow extensively on human milk oligosaccharides as sole carbon source. The genomic sequence of this strain revealed approximately 700 genes that are unique to infantis, including a variety of co-regulated glycosidases, relative

  4. Convergent synthesis of 4,5-branched inner-core oligosaccharides of lipopoly- and lipooligosaccharides.

    PubMed

    Yi, Ruiqin; Narimoto, Hirofumi; Nozoe, Miku; Ichiyanagi, Tsuyoshi

    2015-01-01

    The convergent synthesis of branched inner-core oligosaccharides of lipopoly- and lipooligosaccharide with a 3-deoxy-d-manno-oct-2-ulosonic acid (Kdo) disaccharide acceptor was achieved. The l-glycero-d-manno-heptopyranose (Hep) units for the branched core oligosaccharide Galβ(1-4)Glcβ(1-4)Hep and Hepα(1-3)Hep were prepared from the corresponding Hep building blocks. To obtain 4,5-branched core oligosaccharide structures, the common acceptor Kdoα(2-4)Kdo was glycosylated with the Hep units.

  5. Deacetylation of chitin oligosaccharides of dp 2-4 by chitin deacetylase from Colletotrichum lindemuthianum.

    PubMed

    Tokuyasu, K; Ono, H; Ohnishi-Kameyama, M; Hayashi, K; Mori, Y

    1997-09-26

    Chitin oligosaccharides of degree of polymerization 2-4 were deacetylated by purified chitin deacetylase isolated from Colletotrichum lindemuthianum to give their corresponding breakdown products after purification by liquid chromatography. Data from FABMS analyses suggested that N,N',N",N"'-tetraacetylchitotetraose and N,N',N"-triacetylchitotriose were converted into fully-deacetylated corresponding chitosan oligomers. Conversely, N,N'-diacetylchitobiose [(GlcNAc)2] was deacetylated to give a product which showed an [M + H]+ pseudomolecular ion at m/z 383, suggesting that either of the two acetyl groups were removed. Further data from 1H NMR analyses confirmed that the reaction product was 2-acetamido-4-O-(2-amino-2-deoxy-beta-D-glucopyranosyl)-2-deoxy-D-glucos e [GlcN-GlcNAc]. The enzymatic method has three advantageous characteristics over chemical methods: (i) it does not cause unexpected degradation of the sugar chain, (ii) it is highly reproducible, and (iii) unique compounds such as GlcN-GlcNAc may be produced.

  6. Insights into polymer versus oligosaccharide synthesis: mutagenesis and mechanistic studies of a novel levansucrase from Bacillus megaterium.

    PubMed

    Homann, Arne; Biedendieck, Rebekka; Götze, Sven; Jahn, Dieter; Seibel, Jürgen

    2007-10-15

    A novel levansucrase was identified in the supernatant of a cell culture of Bacillus megaterium DSM319. In order to test for the contribution of specific amino acid residues to levansucrase catalysis, the wild-type enzyme along with 16 variants based on sequence alignments and structural information were heterologously produced in Escherichia coli. The purified enzymes were characterized kinetically and the product spectrum of each variant was determined. Comparison of the X-ray structures of the levansucrases from Gram-positive Bacillus subtilis and Gram-negative Gluconacetobacter diazotrophicus in conjunction with the corresponding product spectra identified crucial amino acid residues responsible for product specificity and catalysis. Highly conserved regions such as the previously described RDP and DXXER motifs were identified as being important. Two crucial structural differences localized at amino acid residues Arg370 and Asn252 were of high relevance in polymer compared with oligosaccharide synthesis.

  7. Structure analysis and laxative effects of oligosaccharides isolated from bananas.

    PubMed

    Wang, Juan; Huang, Hui Hua; Cheng, Yan Feng; Yang, Gong Ming

    2012-10-01

    Banana oligosaccharides (BOS) were extracted with water, and then separated and purified using column chromatography. Gel penetration chromatography was used to determine the molecular weights. Thin layer chromatogram and capillary electrophoresis were employed to analyze the monosaccharide composition. The indican bond and structure of the BOS molecule were determined using Fourier transform infrared spectroscopy and nuclear magnetic resonance. Results showed that BOS were probably composed of eight β-D-pyran glucose units linked with 1→6 indican bonds. The laxative effects of BOS were investigated in mice using the method described in "Handbook of Technical Standards for Testing and Assessment of Health Food in China." The length of the small intestine over which a carbon suspension solution advanced in mice treated with low-, middle-, and high-dose BOS was significantly greater than that in the model group, suggesting that BOS are effective in accelerating the movement of the small intestine.

  8. Plant growth-promoting oligosaccharides produced from tomato waste.

    PubMed

    Suzuki, Toshisada; Tomita-Yokotani, Kaori; Tsubura, Hirokazu; Yoshida, Shigeki; Kusakabe, Isao; Yamada, Kosumi; Miki, Yoichi; Hasegawa, Koji

    2002-01-01

    Tomato juice waste was hydrolyzed with acid. Tomato juice waste (500 g; wet weight) was heated with 0.5 N HCl (2.5 l) at 70 degrees C for 4 h. After neutralization, the growth-promoting extracts (300 g; dry weight) in the plants were produced from the tomato waste. The acid extract significantly promoted the growth of cockscomb (Celosia argentea L.) and tomato (Lycopersicon esculentum L.) seedlings. We have recognized potent plant growth-promoting substances in the acid extract from tomato waste. The most effective components in the active fraction were almost all oligogalacturonic acids (DP 6-12). This paper is the first report that plant growth-promoting oligosaccharides can be directly produced from tomato juice waste. It is possible that the substances from the tomato waste can become useful plant growth regulators in the agriculture field in the future.

  9. A gravimetric analysis of protein-oligosaccharide interactions.

    PubMed

    Rudd, T; Gallagher, J T; Ron, D; Nichols, R J; Fernig, D G

    2003-04-01

    Interactions between an immobilized, heparin-derived octasaccharide and growth factors have been observed using a quartz crystal microbalance-dissipation (QCM-D). This device can measure the amount of growth factors binding to the octasaccharide surface and also the change of dissipation of the surface. Dissipation is a measure of how the adhered material 'damps' the surface vibrations. The octasaccharides were anchored through their reducing ends by the intermediary of the alkanethiol molecule, which covalently binds to the crystal surface through the thiol group. As expected, heparin sulphate binding growth factors bound to the octasaccharide, but the change in mass of growth factor bound per unit change in dissipation is different for the different growth factors. Suggesting that the structures of the various growth factor-octasaccharide complexes are different, therefore, indicates that the change in dissipation can give insights into the structure, orientation and packing of the oligosaccharide-growth factor complexes.

  10. Comparative Genomics of the Aeromonadaceae Core Oligosaccharide Biosynthetic Regions.

    PubMed

    Forn-Cuní, Gabriel; Merino, Susana; Tomás, Juan M

    2017-02-28

    Lipopolysaccharides (LPSs) are an integral part of the Gram-negative outer membrane, playing important organizational and structural roles and taking part in the bacterial infection process. In Aeromonas hydrophila, piscicola, and salmonicida, three different genomic regions taking part in the LPS core oligosaccharide (Core-OS) assembly have been identified, although the characterization of these clusters in most aeromonad species is still lacking. Here, we analyse the conservation of these LPS biosynthesis gene clusters in the all the 170 currently public Aeromonas genomes, including 30 different species, and characterise the structure of a putative common inner Core-OS in the Aeromonadaceae family. We describe three new genomic organizations for the inner Core-OS genomic regions, which were more evolutionary conserved than the outer Core-OS regions, which presented remarkable variability. We report how the degree of conservation of the genes from the inner and outer Core-OS may be indicative of the taxonomic relationship between Aeromonas species.

  11. Characterization of oligosaccharides from an antigenic mannan of Saccharomyces cerevisiae.

    PubMed

    Young, M; Davies, M J; Bailey, D; Gradwell, M J; Smestad-Paulsen, B; Wold, J K; Barnes, R M; Hounsell, E F

    1998-08-01

    Mannans of the yeast Saccharomyces cerevisiae have been implicated as containing the allergens to which bakers and brewers are sensitive and also the antigen recognized by patients with Crohn's disease. A fraction of S. cerevisiae mannan, Sc500, having high affinity for antibodies in Crohn's patients has been characterized by NMR spectroscopy followed by fragmentation using alkaline elimination, partial acid hydrolysis and acetolysis. The released oligosaccharides were separated by gel filtration on a Biogel P4 column and analyzed by fluorescence labeling, HPLC and methylation analysis. The relationship between structure and antigen activity was measured by competitive ELISA. The antigenic activity of the original high molecular weight mannan could be ascribed to terminal Manalpha1-->3Manalpha1-->2 sequences which are rarely found in human glycoproteins but were over-represented in Sc500 compared to other yeast mannans.

  12. Applications of unnatural oligosaccharide biosynthesis to selective drug delivery

    SciTech Connect

    Yarema, K.J.; Mahal, L.K.; Bertozzi, C.R. |

    1997-12-31

    Cell surface antigens are attractive targets for the selective delivery of toxins to diseased cells. Most efforts toward the development of cell-specific drug targeting agents have focused on the use of monoclonal antibodies that recognize a particular antigen, an approach that suffers from severe limitations. We have recently demonstrated that unnatural epitopes can be displayed on cells using the biosynthetic pathways for cell-surface oligosaccharides. This discovery has now been extended to the development of a novel strategy for selective drug targeting. We installed ketone groups on cells through unnatural sialic acid biosynthesis, and selectively modified the cells with biotinamidocaproyl hydrazide under physiological conditions. Synthetic ricin-avidin conjugates killed the biotinylated cells with remarkable selectivity. Applications of this and related approaches to anti-cancer therapy will be presented.

  13. Breast Milk Oligosaccharides: Structure-Function Relationships in the Neonate

    PubMed Central

    Smilowitz, Jennifer T.; Lebrilla, Carlito B.; Mills, David A.; German, J. Bruce; Freeman, Samara L.

    2015-01-01

    In addition to providing complete postnatal nutrition, breast milk is a complex biofluid that delivers bioactive components for the growth and development of the intestinal and immune systems. Lactation is a unique opportunity to understand the role of diet in shaping the intestinal environment including the infant microbiome. Of considerable interest is the diversity and abundance of milk glycans that are energetically costly for the mammary gland to produce yet indigestible by infants. Milk glycans comprise free oligosaccharides, glycoproteins, glycopeptides, and glycolipids. Emerging technological advances are enabling more comprehensive, sensitive, and rapid analyses of these different classes of milk glycans. Understanding the impact of inter- and intraindividual glycan diversity on function is an important step toward interventions aimed at improving health and preventing disease. This review discusses the state of technology for glycan analysis and how specific structure-function knowledge is enhancing our understanding of early nutrition in the neonate. PMID:24850388

  14. Selected microbial glycolipids: production, modification and characterization.

    PubMed

    Palme, Olof; Moszyk, Anja; Iphöfer, Dimitri; Lang, Siegmund

    2010-01-01

    This chapter deals with two types of biosurfactants that are not in the spotlight of general research: glycoglycerolipids and oligosaccharide lipids. The main focus is on glycolglycerolipids from marine bacteria like Microbacterium spec. DSM 12583, Micrococcus luteus (Hel 12/2) and Bacillus pumilus strain AAS3 and on oligosaccharide lipids from Tsukamurella spec. DSM 44370 and Nocardia corynebacteroides SM1. General and special structures, microbial producers, production conditions and chemo-enzymatic modifications as well as properties are outlined.

  15. Recognition of human milk oligosaccharides by bacterial exotoxins.

    PubMed

    El-Hawiet, Amr; Kitova, Elena N; Klassen, John S

    2015-08-01

    The affinities of the most abundant oligosaccharides found in human milk for four bacterial exotoxins (from Vibrio cholerae and pathogenic Escherichia coli) were quantified for the first time. Association constants (Ka) for a library of 20 human milk oligosaccharides (HMOs) binding to Shiga toxin type 2 holotoxin (Stx2) and the B subunit homopentamers of cholera toxin, heat-labile toxin and Shiga toxin type 1 (CTB5, HLTB5 and Stx1B5) were measured at 25°C and pH 7 using the direct electrospray ionization mass spectrometry assay. Notably, all four bacterial toxins bind to a majority of the HMOs tested and five of the HMOs (2'-fucosyllactose, lacto-N-tetraose, lacto-N-fucopentaose I, lacto-N-fucopentaose II and lacto-N-fucopentaose III) are ligands for all four toxins. These five HMOs are also reported to bind to other bacterial toxins (e.g. toxin A and toxin B of Clostridium difficile). In all cases, the HMO affinities (apparent Ka) are relatively modest (≤15,000 M(-1)). However, at the high concentrations of HMOs typically ingested by infants, a significant fraction of these toxins, if present, is expected to be bound to HMOs. Binding measurements carried out with 2'-fucosyllactose or lacto-N-fucopentaose I, together with a high-affinity ligand based on the native carbohydrate receptor, revealed that all four toxins possess HMO-binding sites that are distinct from those of the native receptors, although evidence of competitive binding was found for lacto-N-fucopentaose I with Stx2 and 2'-fucosyllactose and lacto-N-fucopentaose I with HLTB5. Taken together, the results of this study suggest that, while HMOs are expected to bind extensively to these bacterial toxins, it is unlikely that HMO binding will effectively inhibit their interactions with their cellular receptors.

  16. Human milk oligosaccharides: every baby needs a sugar mama.

    PubMed

    Bode, Lars

    2012-09-01

    Human milk oligosaccharides (HMOs) are a family of structurally diverse unconjugated glycans that are highly abundant in and unique to human milk. Originally, HMOs were discovered as a prebiotic "bifidus factor" that serves as a metabolic substrate for desired bacteria and shapes an intestinal microbiota composition with health benefits for the breast-fed neonate. Today, HMOs are known to be more than just "food for bugs". An accumulating body of evidence suggests that HMOs are antiadhesive antimicrobials that serve as soluble decoy receptors, prevent pathogen attachment to infant mucosal surfaces and lower the risk for viral, bacterial and protozoan parasite infections. In addition, HMOs may modulate epithelial and immune cell responses, reduce excessive mucosal leukocyte infiltration and activation, lower the risk for necrotizing enterocolitis and provide the infant with sialic acid as a potentially essential nutrient for brain development and cognition. Most data, however, stem from in vitro, ex vivo or animal studies and occasionally from association studies in mother-infant cohorts. Powered, randomized and controlled intervention studies will be needed to confirm relevance for human neonates. The first part of this review introduces the pioneers in HMO research, outlines HMO structural diversity and describes what is known about HMO biosynthesis in the mother's mammary gland and their metabolism in the breast-fed infant. The second part highlights the postulated beneficial effects of HMO for the breast-fed neonate, compares HMOs with oligosaccharides in the milk of other mammals and in infant formula and summarizes the current roadblocks and future opportunities for HMO research.

  17. Human milk oligosaccharides: Every baby needs a sugar mama

    PubMed Central

    Bode, Lars

    2012-01-01

    Human milk oligosaccharides (HMOs) are a family of structurally diverse unconjugated glycans that are highly abundant in and unique to human milk. Originally, HMOs were discovered as a prebiotic “bifidus factor” that serves as a metabolic substrate for desired bacteria and shapes an intestinal microbiota composition with health benefits for the breast-fed neonate. Today, HMOs are known to be more than just “food for bugs”. An accumulating body of evidence suggests that HMOs are antiadhesive antimicrobials that serve as soluble decoy receptors, prevent pathogen attachment to infant mucosal surfaces and lower the risk for viral, bacterial and protozoan parasite infections. In addition, HMOs may modulate epithelial and immune cell responses, reduce excessive mucosal leukocyte infiltration and activation, lower the risk for necrotizing enterocolitis and provide the infant with sialic acid as a potentially essential nutrient for brain development and cognition. Most data, however, stem from in vitro, ex vivo or animal studies and occasionally from association studies in mother–infant cohorts. Powered, randomized and controlled intervention studies will be needed to confirm relevance for human neonates. The first part of this review introduces the pioneers in HMO research, outlines HMO structural diversity and describes what is known about HMO biosynthesis in the mother's mammary gland and their metabolism in the breast-fed infant. The second part highlights the postulated beneficial effects of HMO for the breast-fed neonate, compares HMOs with oligosaccharides in the milk of other mammals and in infant formula and summarizes the current roadblocks and future opportunities for HMO research. PMID:22513036

  18. Kinetic characterization of Aspergillus niger chitinase CfcI using a HPAEC-PAD method for native chitin oligosaccharides.

    PubMed

    van Munster, Jolanda M; Sanders, Peter; ten Kate, Geralt A; Dijkhuizen, Lubbert; van der Maarel, Marc J E C

    2015-04-30

    The abundant polymer chitin can be degraded by chitinases (EC 3.2.1.14) and β-N-acetyl-hexosaminidases (EC 3.2.1.52) to oligosaccharides and N-acetyl-glucosamine (GlcNAc) monomers. Kinetic characterization of these enzymes requires product quantification by an assay method with a low detection limit, preferably compatible with the use of native, non-labeled substrates. Here we report a quantitative HPAEC-PAD method that allows fast separation of chitin oligosaccharides (COS) ranging from (GlcNac)1-6 at detection limits of 1-3 pmol and a linear range of 5-250 pmol. Quantification under intra- and interday precision conditions was performed with 2.1-5.4% relative standard deviation (RSD) and 1.2-10.3% RSD, respectively. This method was successfully used for the determination of the kinetic parameters of the Aspergillus niger chitinase CfcI with native COS. CfcI was recently shown to release GlcNAc from the reducing end of COS, a new activity for fungal chitinases. A Carbohydrate Binding Module of family 18 (CBM18) is inserted in the CfcI catalytic domain. Site directed mutagenesis was used to assess the functionality of this CfcI-CBM18: four of its key amino acids were replaced by glycine residues, yielding CfcISYNF. Comparison of the kinetic parameters of CfcI and CfcISYNF confirmed that this CBM18 is functionally involved in catalysis.

  19. Isolation and characterization of feruloylated arabinoxylan oligosaccharides from the perennial cereal grain intermediate wheat grass (Thinopyrum intermedium).

    PubMed

    Schendel, Rachel R; Becker, Andreas; Tyl, Catrin E; Bunzel, Mirko

    2015-04-30

    In comparison to the annual grain crops dominating current agricultural production, perennial grain species require fewer chemical and energy inputs and improve soil health and erosion control. The possibility for producing sustainable grain harvests from marginal land areas is motivating research initiatives to integrate perennial grains into commercial cropping and food processing systems. In this study, the feruloylated arabinoxylans from intermediate wheat grass (Thinopyrum intermedium, IWG), a promising perennial grain candidate in agronomic screening studies, were investigated. Insoluble fiber isolated from IWG whole grain flour was subjected to either mildly acidic (50 mM TFA, 100 °C, 2 h) or enzymatic (Driselase) hydrolysis. The liberated feruloylated arabinoxylan oligosaccharides were concentrated with Amberlite XAD-2, separated with gel chromatography (Sephadex LH-20, water), and purified with reversed-phase HPLC (C18, water-MeOH gradient). Thirteen feruloylated oligosaccharides were isolated (including eight structures described for the first time) and identified by LC-ESI-MS and NMR. Linkage-type analysis via methylation analysis, as well as the monosaccharide and phenolic acid profiles of the IWG insoluble fiber were also determined. IWG feruloylated arabinoxylans have a relatively simple structure with only short feruloylated side chains, a lower backbone substitution rate than annual rye and wheat varieties, and a moderate phenolic acid content. Copyright © 2015 Elsevier Ltd. All rights reserved.

  20. Different effects of native Candida albicans mannan and mannan-derived oligosaccharides on antigen-stimulated lymphoproliferation in vitro.

    PubMed

    Podzorski, R P; Gray, G R; Nelson, R D

    1990-01-15

    Yeast cell wall mannan polysaccharide has been proposed to contribute to immune dysfunction associated with chronic infections involving Candida albicans. This influence of mannan has been suggested based partially upon studies of the in vitro immunoinhibitory effects of mannans prepared from Saccharomyces cerevisiae or C. albicans by precipitation with Fehling's reagent, which provides a structurally modified product contaminated with copper. We have therefore evaluated the immunoinhibitory influence of a more native C. albicans mannan prepared by complexation with cetyltrimethylammonium bromide (CTAB). CTAB mannan was a potent stimulator of lymphoproliferation when added to human PMBC from donors responsive to Candida; it has no inhibitory influence on lympho-proliferation induced by Candida or other Ag. In contrast, members of a family of mannose oligosaccharides (disaccharide through hexasaccharide) derived from the CTAB mannan by weak alkaline degradation did not stimulate lymphoproliferation, but were potent inhibitors of lymphoproliferation stimulated by Candida and other Ag. Fifty percent inhibitory doses were 260 to 34 microM, respectively. Compositional analyses of these immunoinhibitory oligomers showed them to be composed of mannose and free of contaminating protein. We propose that mannan-derived oligosaccharides produced by catabolism of mannan in vivo are immunoinhibitory and contribute to the deficit in cell-mediated immune function associated with chronic candidiasis.

  1. Milk oligosaccharide sialyl(α2,3)lactose activates intestinal CD11c+ cells through TLR4.

    PubMed

    Kurakevich, Ekaterina; Hennet, Thierry; Hausmann, Martin; Rogler, Gerhard; Borsig, Lubor

    2013-10-22

    Breast milk oligosaccharides shape the intestinal environment by affecting mucosal immunity and bacterial colonization. To clarify the role of milk oligosaccharide sialyl(α2,3)lactose (3SL) in intestinal physiology and disease, we investigated colitis development in Il10(-/-) mice exposed to normal or 3SL-deficient milk during lactation. Onset and progression of intestinal inflammation were delayed in Il10(-/-) mice deficient for the α2,3 sialyltransferase 4 (ST3GAL4) responsible for 3SL biosynthesis. The proinflammatory role of 3SL was confirmed by showing that oral supplementation of newborn Il10(-/-);St3gal4(-/-) mice with 3SL increased colitis severity. Conversely, fostering of newborn Il10(-/-) mice to lactating St3gal4(-/-) mothers reduced colitis severity. 3SL directly stimulated mesenteric lymph node CD11c(+) dendritic cells and induced production of cytokines required for expansion of TH1 and TH17 T cells. The stimulatory effect of 3SL was attenuated in Tlr4-deficient CD11c(+) cells, demonstrating that 3SL induces inflammation through Toll-like receptor 4 (TLR4) signaling. Thus, 3SL directly modulates mucosal immunity, which increases susceptibility to colitis.

  2. Capillary electrophoresis separation of a mixture of chitin and chitosan oligosaccharides derivatized using a modified fluorophore conjugation procedure.

    PubMed

    Beaudoin, Marie-Eve; Gauthier, Julie; Boucher, Isabelle; Waldron, Karen C

    2005-08-01

    A capillary electrophoresis (CE) method was developed for the simultaneous analysis of small chitin and chitosan oligosaccharides. For detection purposes, the oligomers were derivatized with 8-aminopyrene-1,3,6-trisulfonic acid (APTS), a well known fluorophore for oligosaccharides analysis. The detection was performed by laser-induced fluorescence (LIF) with an argon ion laser having an excitation wavelength of 488 nm and with emission monitored at 520 nm. Derivatization parameters such as reaction time and conditions were examined. Separation conditions were also varied by testing a range of buffer pHs and concentrations. The best conditions were found using an 80 mM borate buffer at pH 8.4. This CE-LIF optimized method was used for the analysis of an enzymatically produced oligo-chitosan sample composed of a complex mixture and having an average degree of polymerization of 3.7 monomer units and 80% deacetylation. The oligo-chitosan sample was treated with a chitin deacetylase-like enzyme, the products were derivatized with APTS, and then analyzed without purification. The goal was to determine whether the deacetylase-like enzyme could increase the extent of deacetylation of the oligo-chitosan sample.

  3. Regulation of the protein glycosylation pathway in yeast: structural control of N-linked oligosaccharide elongation

    SciTech Connect

    Gopal, P.K.; Ballou, C.E.

    1987-12-01

    The yeast Saccharomyces cerevisiae X2180 strain with the mnn1 mnn2 mnn9 mutations, all of which affect mannoprotein glycosylation, synthesizes N-linked oligosaccharides. Membrane fractions from the mnn1 mnn2 and mnn1 mnn2 mnn9 mutants are equally effective in catalyzing transfer from GDP-(/sup 3/H)mannose to add mannose in both ..cap alpha..1 ..-->.. 2 and ..cap alpha..1 ..-->.. 6 linkages to an oligosaccharide. Neither membrane preparation can utilize the homologous mnn1 mnn2 mnn9 oligosaccharide as an acceptor. Thus, addition of the ..cap alpha..1 ..-->.. 2-linked mannose side chain to the terminal ..cap alpha..1 ..-->.. 6-linked mannose in oligosaccharides of the mnn9 mutant inhibits the elongation reaction and may serve as an important structural control of mannoprotein glycosylation. The mnn9 mutation also increases the transit time for invertase secretion, meaning that this mutation could affect the processing machinery in the Golgi apparatus.

  4. (Pseudo)amide-linked oligosaccharide mimetics: molecular recognition and supramolecular properties

    PubMed Central

    Ortega-Caballero, Fernando; Ortiz Mellet, Carmen; García Fernández, José M

    2010-01-01

    Summary Oligosaccharides are currently recognised as having functions that influence the entire spectrum of cell activities. However, a distinct disadvantage of naturally occurring oligosaccharides is their metabolic instability in biological systems. Therefore, much effort has been spent in the past two decades on the development of feasible routes to carbohydrate mimetics which can compete with their O-glycosidic counterparts in cell surface adhesion, inhibit carbohydrate processing enzymes, and interfere in the biosynthesis of specific cell surface carbohydrates. Such oligosaccharide mimetics are potential therapeutic agents against HIV and other infections, against cancer, diabetes and other metabolic diseases. An efficient strategy to access this type of compounds is the replacement of the glycosidic linkage by amide or pseudoamide functions such as thiourea, urea and guanidine. In this review we summarise the advances over the last decade in the synthesis of oligosaccharide mimetics that possess amide and pseudoamide linkages, as well as studies focussing on their supramolecular and recognition properties. PMID:20485602

  5. Preparation and antioxidant activity of the oligosaccharides derived from Laminaria japonica.

    PubMed

    Wu, Sheng-Jun

    2014-06-15

    Hydrogen peroxide (H2O2) can degrade polysaccharides and has bleaching effect. In this study, the oligosaccharides derived from Laminaria japonica were prepared by hydrolysis with H2O2 and their antioxidant activity was investigated. The optimal hydrolysis conditions were determined as follows: reaction time 24h, reaction temperature 75°C, and H2O2 concentration 4%. Under the optimum conditions, the maximum yield of the oligosaccharides reached 17.65%, which was higher than that of aqueous extraction, and at the same time, the maximal decoloration rate reached 79.85%. The oligosaccharides sample contained 94.82% sugar, of which the average degree was approximately 8, and showed light green. The oligosaccharides derived from L. japonica showed high hydroxyl radical scavenging activity (91.31%) at the concentration of 100 μg/mL. Copyright © 2014 Elsevier Ltd. All rights reserved.

  6. The hydrolyzation of collagen by fucoidan oligosaccharide's complex with CeIV

    NASA Astrophysics Data System (ADS)

    Xu, Jiachao; Gao, Xin; Zhang, Zhaohui; Li, Zhaojie; Huo, Lihua; Xue, Changhu

    2006-04-01

    Fucoidan is such a polysaccharide that its hydroxies are easy to combine with lanthanons ion (CeIV) to form complex. This work obtained the complexes of three fucoidan oligosaccharides with different molecular weights F1 (>5000), F2 (1000 5000) and F3 (<1000) by hydrolyzing Oligosaccharide collagen with sulfuric acid. It is found that the fucoidan oligosaccharide F3 can form complex with more CeIV than F1 and F2. Hydrolyzing collagen with the complex was carried out to produce amino acid and peptides. All the three fucoidan oligosaccharide complexes with CeIV (F1, F2, F3) can catalyze by the artificial hydrolytic enzyme, and the activity of the complex of F3 is the highest.

  7. Elucidation of the structure of the oligosaccharide from wild type Moraxella bovis Epp63 lipooligosaccharide.

    PubMed

    De Castro, Cristina; Grice, I Darren; Daal, Terese-Marie; Peak, Ian R; Molinaro, Antonio; Wilson, Jennifer C

    2014-03-31

    Moraxella bovis is a Gram-negative microorganism that causes Infectious Bovine Keratoconjunctivitis (IBK), colloquially known as 'Pink eye' in cattle worldwide. Lipopolysaccharides/lipooligosaccharides are the predominant glycans on the surface of Gram-negative microorganisms. Structural elucidation of the oligosaccharide structure of the rough phenotype of Moraxella bovis strain Epp63 was determined using GC-MS, methylation analysis, and NMR spectroscopy. The oligosaccharide is a branched structure that comprises 10 sugars in addition to KDO. The unusual features of this oligosaccharide include the fact that the oligosaccharide is devoid of heptose. The KDO residue is directly attached to a (→4,6)-branched glucose and additionally contains a terminal open chain acetal-linked N-acetylgalactosamine, (1S)-GalaNAc residue →4,6-linked to a sub-terminal galactose residue.

  8. Stability of oligosaccharides derived from lactulose during the processing of milk and apple juice.

    PubMed

    López-Sanz, Sara; Montilla, Antonia; Moreno, F Javier; Villamiel, Mar

    2015-09-15

    The scientific evidence on the bioactivity of oligosaccharides from lactulose has encouraged us to study their physicochemical modifications during the processing of milk and apple juice. The carbohydrate fraction with a degree of polymerization ⩾3 was stable in milk heated at temperatures up to 100°C for 30min and in apple juice heated up to 90°C for 15min. An assessment of the Maillard reaction in heated milk pointed out a higher formation of furosine in milk with oligosaccharides from lactulose as compared to its counterpart without this ingredient, due to a higher presence of galactose. The organoleptic properties of juice with oligosaccharides from lactulose were acceptable and similar to those of apple juice with commercial galactooligosaccharides. The results presented herein demonstrate that oligosaccharides from lactulose can be used as prebiotic ingredients in a wide range of functional foods, including those intended for diabetics and lactose intolerant individuals. Copyright © 2015 Elsevier Ltd. All rights reserved.

  9. Linkage Determination of Linear Oligosaccharides by MSn (n > 2) Collision-Induced Dissociation of Z1 Ions in the Negative Ion Mode

    NASA Astrophysics Data System (ADS)

    Konda, Chiharu; Bendiak, Brad; Xia, Yu

    2013-12-01

    Obtaining unambiguous linkage information between sugars in oligosaccharides is an important step in their detailed structural analysis. An approach is described that provides greater confidence in linkage determination for linear oligosaccharides based on multiple-stage tandem mass spectrometry (MSn, n >2) and collision-induced dissociation (CID) of Z1 ions in the negative ion mode. Under low energy CID conditions, disaccharides 18O-labeled on the reducing carbonyl group gave rise to Z1 product ions (m/z 163) derived from the reducing sugar, which could be mass-discriminated from other possible structural isomers having m/z 161. MS3 CID of these m/z 163 ions showed distinct fragmentation fingerprints corresponding to the linkage types and largely unaffected by sugar unit identities or their anomeric configurations. This unique property allowed standard CID spectra of Z1 ions to be generated from a small set of disaccharide samples that were representative of many other possible isomeric structures. With the use of MSn CID (n = 3 - 5), model linear oligosaccharides were dissociated into overlapping disaccharide structures, which were subsequently fragmented to form their corresponding Z1 ions. CID data of these Z1 ions were collected and compared with the standard database of Z1 ion CID using spectra similarity scores for linkage determination. As the proof-of-principle tests demonstrated, we achieved correct determination of individual linkage types along with their locations within two trisaccharides and a pentasaccharide.

  10. Application of Metabolic 13C Labeling in Conjunction with High-Field Nuclear Magnetic Resonance Spectroscopy for Comparative Conformational Analysis of High Mannose-Type Oligosaccharides

    PubMed Central

    Kamiya, Yukiko; Yanagi, Kotaro; Kitajima, Toshihiko; Yamaguchi, Takumi; Chiba, Yasunori; Kato, Koichi

    2013-01-01

    High mannose-type oligosaccharides are enzymatically trimmed in the endoplasmic reticulum, resulting in various processing intermediates with exposed glycotopes that are recognized by a series of lectins involved in glycoprotein fate determination in cells. Although recent crystallographic data have provided the structural basis for the carbohydrate recognition of intracellular lectins, atomic information of dynamic oligosaccharide conformations is essential for a quantitative understanding of the energetics of carbohydrate–lectin interactions. Carbohydrate NMR spectroscopy is useful for characterizing such conformational dynamics, but often hampered by poor spectral resolution and lack of recombinant techniques required to produce homogeneous glycoforms. To overcome these difficulties, we have recently developed a methodology for the preparation of a homogeneous high mannose-type oligosaccharide with 13C labeling using a genetically engineered yeast strain. We herein successfully extended this method to result in the overexpression of 13C-labeled Man9GlcNAc2 (M9) with a newly engineered yeast strain with the deletion of four genes involved in N-glycan processing. This enabled high-field NMR analyses of 13C-labeled M9 in comparison with its processing product lacking the terminal mannose residue ManD2. Long-range NOE data indicated that the outer branches interact with the core in both glycoforms, and such foldback conformations are enhanced upon the removal of ManD2. The observed conformational variabilities might be significantly associated with lectins and glycan-trimming enzymes. PMID:24970159

  11. Linkage determination of linear oligosaccharides by MS(n) (n > 2) collision-induced dissociation of Z₁ ions in the negative ion mode.

    PubMed

    Konda, Chiharu; Bendiak, Brad; Xia, Yu

    2014-02-01

    Obtaining unambiguous linkage information between sugars in oligosaccharides is an important step in their detailed structural analysis. An approach is described that provides greater confidence in linkage determination for linear oligosaccharides based on multiple-stage tandem mass spectrometry (MS(n), n >2) and collision-induced dissociation (CID) of Z1 ions in the negative ion mode. Under low energy CID conditions, disaccharides (18)O-labeled on the reducing carbonyl group gave rise to Z1 product ions (m/z 163) derived from the reducing sugar, which could be mass-discriminated from other possible structural isomers having m/z 161. MS(3) CID of these m/z 163 ions showed distinct fragmentation fingerprints corresponding to the linkage types and largely unaffected by sugar unit identities or their anomeric configurations. This unique property allowed standard CID spectra of Z1 ions to be generated from a small set of disaccharide samples that were representative of many other possible isomeric structures. With the use of MS(n) CID (n = 3 - 5), model linear oligosaccharides were dissociated into overlapping disaccharide structures, which were subsequently fragmented to form their corresponding Z1 ions. CID data of these Z1 ions were collected and compared with the standard database of Z1 ion CID using spectra similarity scores for linkage determination. As the proof-of-principle tests demonstrated, we achieved correct determination of individual linkage types along with their locations within two trisaccharides and a pentasaccharide.

  12. Comparative study of blood group-recognizing lectins toward ABO blood group antigens on neoglycoproteins, glycoproteins and complex-type oligosaccharides.

    PubMed

    Matsui, T; Hamako, J; Ozeki, Y; Titani, K

    2001-02-16

    Binding specificities of ABO blood group-recognizing lectins toward blood group antigens on neoglycoproteins, glycoproteins and complex-type oligosaccharides were studied by lectin-blotting analysis, enzyme linked immunosorbent assay and lectin-conjugated agarose column chromatography. Human serum albumin conjugated with A- and B-trisaccharides was clearly recognized by Helix pomatia (HPA), Phaseolus lunatus, Dolichos biflorus agglutinins, and Griffonia simplicifolia I agglutinin B(4), respectively. Almost the same results were obtained for human group A and B ovarian cyst and A-active hog gastric mucins, but Glycine max agglutinin only reacted to the group A hog mucin. When human plasma von Willebrand factor (vWF), having Asn-linked blood group antigens, was tested, HPA was highly sensitive to blood group A antigen on the vWF. Ulex europaeus agglutinin I (UEA-I) preferentially bound to the vWF from blood group O plasma. Within the GalNAc-recognizing lectins examined, a biantennary complex-type oligosaccharide having the blood group A structure retarded on an HPA-agarose column, and the affinity was diminished after digestion with alpha-N-acetylgalactosaminidase. This product bound to UEA-I agarose column. These results indicate that HPA and UEA-I are most sensitive for detection of glycoproteins possessing small amounts of blood group A and H antigens and also useful for fractionation of complex-type oligosaccharides with blood group A and H antigens, respectively.

  13. Synthetic glycosylation of proteins using N-(beta-saccharide) iodoacetamides: applications in site-specific glycosylation and solid-phase enzymic oligosaccharide synthesis.

    PubMed Central

    Wong, S Y; Guile, G R; Dwek, R A; Arsequell, G

    1994-01-01

    A simple and efficient synthetic glycosylation method suitable for use in solid-phase enzymic oligosaccharide synthesis and site-specific glycosylation of recombinant proteins to produce defined glycoforms is described. This strategy utilizes N-(beta-saccharide) haloacetamides for attaching oligosaccharides specifically to cysteine residues of proteins in solution to form neoglycoproteins. The alkylation reaction was tested using N-(beta-chitotriose) bromoacetamide and an unprotected synthetic hexapeptide containing a single cysteine residue. The glycosylated product was confirmed by amino acid and hexosamine analyses as well as laser desorption mass spectrometry. Similarly N-(beta-chitotriose) iodoacetamide was covalently linked to non-reduced BSA to produce a defined glycoform of this protein. The specific attachment of chitotriose at the single cysteine residue in non-reduced serum albumin was suggested by Ellman's assay for free thiols. This was verified by amino acid sequencing of tryptic glycopeptide derived from this neoglycoprotein. Multiple sugar attachment was accomplished using fully reduced serum albumin as demonstrated by the formation of two neoglycoproteins using iodoacetamide derivatives of galactose beta 1-3-N-acetylgalactosamine (Gal beta 1-3GalNAc) and the major xylose/fucose-class plant-type oligosaccharide of horseradish peroxidase. These two neoglycoproteins with an average of 18-21 sugar residues attached were assayed positively for binding to peanut agglutinin and a sugar-specific anti-(horseradish peroxidase) monoclonal antibody YZ1/2.23 respectively. Sialylation of the neoglycoprotein containing Gal beta 1-3GalNAc was accomplished using alpha-2,3-sialyltransferase and radiolabelled CMP-N-acetylneuraminic acid. Significantly, glycan attachment using this conjugation method is reversible as demonstrated by the release of oligosaccharides from these two neoglycoproteins using hydrazinolysis. Therefore this method could provide invaluable

  14. Effects of chito-oligosaccharides supplementation on growth performance, intestinal cytokine expression, autochthonous gut bacteria and disease resistance in hybrid tilapia Oreochromis niloticus ♀ × Oreochromis aureus ♂.

    PubMed

    Qin, Chubin; Zhang, Yuting; Liu, Wenshu; Xu, Li; Yang, Yalin; Zhou, Zhigang

    2014-09-01

    We investigated the effects of incorporating chitinase (ChiB565)-hydrolyzed shrimp shell chitin into the diet of hybrid tilapia (Oreochromis niloticus ♀ × Oreochromis aureus ♂) with regard to production, intestinal immune status and autochthonous gut bacteria, and protection against bacterial pathogen Aeromonas hydrophila. Five experimental diets were formulated by supplementing the basal diet with the hydrolyzed shrimp shell chitin (0.0%, T1 control; 0.8%, T3; 1.6%, T4; or 2.4%, T5) or 0.1% commercial chitosan-oligosaccharides as commercial recommendation dose (T2, positive control). After a 35-day feeding trial, we found no significant difference in weight gain, feed conversion ratio or survival rate in tilapia among all treatment groups. However, the levels of mRNAs encoding the pro-inflammatory protein tumor necrosis factor-α and the stress-response protein heat shock protein 70 were much lower in groups T2, T3, T4 and T5 (p < 0.001). The levels of transforming growth factor-β were higher in groups T2 and T4 (p < 0.001 and p < 0.0001, respectively). In addition, group T3 and T4 with 0.8% and 1.6% hydrolyzed shrimp shell chitin supplementation respectively changed marginally their autochthonous gut bacteria (0.60 < Cs < 0.80). When challenged with A. hydrophila, the mortality of groups fed chito-oligosaccharides was lower than the control, especially in groups T4 and T5 (p < 0.05). These results indicate that dietary intake of chito-oligosaccharides can improve intestinal health, changed autochthonous gut bacteria, and improve resistance to infection by A. hydrophila, even with higher efficiency than receiving the manufacturer recommended dose of the commercial chitosan-oligosaccharides.

  15. Synthetic glycosylation of proteins using N-(beta-saccharide) iodoacetamides: applications in site-specific glycosylation and solid-phase enzymic oligosaccharide synthesis.

    PubMed

    Wong, S Y; Guile, G R; Dwek, R A; Arsequell, G

    1994-06-15

    A simple and efficient synthetic glycosylation method suitable for use in solid-phase enzymic oligosaccharide synthesis and site-specific glycosylation of recombinant proteins to produce defined glycoforms is described. This strategy utilizes N-(beta-saccharide) haloacetamides for attaching oligosaccharides specifically to cysteine residues of proteins in solution to form neoglycoproteins. The alkylation reaction was tested using N-(beta-chitotriose) bromoacetamide and an unprotected synthetic hexapeptide containing a single cysteine residue. The glycosylated product was confirmed by amino acid and hexosamine analyses as well as laser desorption mass spectrometry. Similarly N-(beta-chitotriose) iodoacetamide was covalently linked to non-reduced BSA to produce a defined glycoform of this protein. The specific attachment of chitotriose at the single cysteine residue in non-reduced serum albumin was suggested by Ellman's assay for free thiols. This was verified by amino acid sequencing of tryptic glycopeptide derived from this neoglycoprotein. Multiple sugar attachment was accomplished using fully reduced serum albumin as demonstrated by the formation of two neoglycoproteins using iodoacetamide derivatives of galactose beta 1-3-N-acetylgalactosamine (Gal beta 1-3GalNAc) and the major xylose/fucose-class plant-type oligosaccharide of horseradish peroxidase. These two neoglycoproteins with an average of 18-21 sugar residues attached were assayed positively for binding to peanut agglutinin and a sugar-specific anti-(horseradish peroxidase) monoclonal antibody YZ1/2.23 respectively. Sialylation of the neoglycoprotein containing Gal beta 1-3GalNAc was accomplished using alpha-2,3-sialyltransferase and radiolabelled CMP-N-acetylneuraminic acid. Significantly, glycan attachment using this conjugation method is reversible as demonstrated by the release of oligosaccharides from these two neoglycoproteins using hydrazinolysis. Therefore this method could provide invaluable

  16. Effects of Xylo-Oligosaccharides on Broiler Chicken Performance and Microbiota

    PubMed Central

    De Maesschalck, C.; Eeckhaut, V.; Maertens, L.; De Lange, L.; Marchal, L.; Nezer, C.; De Baere, S.; Croubels, S.; Daube, G.; Dewulf, J.; Haesebrouck, F.; Ducatelle, R.; Taminau, B.

    2015-01-01

    In broiler chickens, feed additives, including prebiotics, are widely used to improve gut health and to stimulate performance. Xylo-oligosaccharides (XOS) are hydrolytic degradation products of arabinoxylans that can be fermented by the gut microbiota. In the current study, we aimed to analyze the prebiotic properties of XOS when added to the broiler diet. Administration of XOS to chickens, in addition to a wheat-rye-based diet, significantly improved the feed conversion ratio. XOS significantly increased villus length in the ileum. It also significantly increased numbers of lactobacilli in the colon and Clostridium cluster XIVa in the ceca. Moreover, the number of gene copies encoding the key bacterial enzyme for butyrate production, butyryl-coenzyme A (butyryl-CoA):acetate CoA transferase, was significantly increased in the ceca of chickens administered XOS. In this group of chickens, at the species level, Lactobacillus crispatus and Anaerostipes butyraticus were significantly increased in abundance in the colon and cecum, respectively. In vitro fermentation of XOS revealed cross-feeding between L. crispatus and A. butyraticus. Lactate, produced by L. crispatus during XOS fermentation, was utilized by the butyrate-producing Anaerostipes species. These data show the beneficial effects of XOS on broiler performance when added to the feed, which potentially can be explained by stimulation of butyrate-producing bacteria through cross-feeding of lactate and subsequent effects of butyrate on gastrointestinal function. PMID:26092452

  17. Oligosaccharide structure determines prebiotic role of β-galactomannan against Salmonella enterica ser. Typhimurium in vitro

    PubMed Central

    Badia, Roger; Lizardo, Rosil; Martínez, Paz; Brufau, Joaquim

    2013-01-01

    Prebiotics and probiotics are considered natural alternatives to dietary antibiotics in animal production. Plant extracts and yeast cell walls are mannose—rich products that can be used as substrate for adhesion of Gram-negative bacteria. We assessed whether the structure of these saccharides is relevant to develop their role as prebiotics and therefore, their suitability to be used as alternatives to antibiotics to prevent intestinal infections in pigs. The prebiotic functionality of β-galactomannan (βGM), mannanoligosaccharide from yeast Saccharomyces cerevisiae (Mannan SC) and monosaccharide D-Mannose were studied in porcine intestinal epithelial cells (IPI-2I) challenged with Salmonella enterica ser. Typhimurium. Results sh