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Ulvan, a sulfated polysaccharide extracted from the green seaweed genus Ulva, has bioactive properties including an immunomodulating capacity. The immunomodulatory capacity of ulvan from Ulva ohnoi, however, has not been assessed in detail. We depolymerised purified ulvan from U. ohnoi to obtain a range of molecular weight fractions (Mw 7, 9, 13, 21, 209 kDa), which were characterised by constituent sugar analysis, SEC-MALLS, and NMR. Ulvan fractions contained 48.8–54.7 mol% rhamnose, 32.5–35.9 mol% glucuronic acid, 4.5–7.3 mol% iduronic acid, and 3.3–5.6 mol% xylose. 1H and 13C NMR was consistent with hydrolysis occurring at the anomeric centre without further modification to the oligosaccharide structure. The in vitro immunomodulatory effect of ulvan fractions was quantified by measuring levels of inflammatory-mediating signalling molecules released from LPS-stimulated RAW264.7 murine macrophages. All ulvan fractions showed no toxicity on RAW264.7 cells at concentrations below 100 μg mL−1 over 48 h. Secreted interleukin-10 and prostaglandin E2 demonstrated an anti-inflammatory effect by higher molecular weight ulvan fractions at 100 μg mL−1. To a lesser extent, these fractions also enhanced the LPS-induced inflammation through minor increases of IL-1β and IL-6. This study confirms that ulvan from U. ohnoi has a mild in vitro immunomodulatory effect.

Background Human health depends on a population of microorganisms that inhabit the gut and contribute to homeostasis of the host, including nutrition, immunity and metabolism. Many of the organisms are interactive and mutually dependent, where the end-products for one organism become the fuel for another through substrate and metabolic cross-feeding. To optimise the gut microbiota using diet, the composition and functionality of the gut microbiota, including these interacting networks, must be understood. Microbial composition and functionality is affected by the structure of the energy input, which is primarily dietary fibre for the gut microbiota. The structure of dietary fibre has been reviewed by carbohydrate chemists, but knowledge of how dietary fibre structure affects the gut microbiota is limited. Scope and approach The hierarchical structures of dietary fibre are reviewed, encompassing macrostructure, mesostructure and molecular structure, and how they are affected by food processing and digestion. These factors are considered in relation to their affects on microbial composition and functionality, to provide insight on the interactions between diet, the microbiota, and human health. Key findings and conclusions Food processing and digestion affect food structure, primarily through the removal of some soluble fractions and increased solubilisation of insoluble fractions. The provision of insoluble carbohydrates to the colon appears important for the sustenance of ‘keystone’ species that play a crucial role in stabilising the gut community. Further work is needed at the microbial strain level to understand the impact of increasing fibre solubility. This should be done in studies using well-characterised carbohydrates that consider the impact of food processing and digestion.

A water-soluble polysaccharide (type II arabinogalactan-protein) extracted from the gum exudate of the native New Zealand puka tree (Meryta sinclairii), was characterised for its molecular, rheological and physicochemical properties. In 0.1 M NaCl, the weight average molecular weight (Mw) of puka gum is 5.9 × 106 Da with an RMS radius of 56 nm and z-average hydrodynamic radius of 79 nm. The intrinsic viscosity of the polysaccharide is 57 ml/g with a coil overlap concentration 15% w/w. Together, the shape factor, p, of 0.70 (exponent of RMS radius vs. hydrodynamic radius), Smidsrod-Haug’s stiffness parameter B of 0.031 and Mark-Houwink exponent α of 0.375 indicate that the polysaccharide adopts a spherical conformation in solution, similar to gum arabic. The pKa is 1.8. The polysaccharide exhibits a Newtonian to shear-thinning behaviour from 0.2 to 25% w/w. Viscosity of the polysaccharide (1 s−1) decreases with decreasing concentration, increasing temperature, ionic strength, and at acidic pH.

Commercial porcine intestinal mucosal heparan sulfate (HS) is a valuable material for research into its biological functions. As it is usually produced as a side-stream of pharmaceutical heparin manufacture, its chemical composition may vary from batch to batch. We analysed the composition and structure of nine batches of HS from the same manufacturer. Statistical analysis of the disaccharide compositions placed these batches in three categories: group A had high GlcNAc and GlcNS, and low GlcN typical of HS; group B had high GlcN and GlcNS, and low GlcNAc; group C had high di- and trisulfated, and low unsulfated and monosulfated disaccharide repeats. These batches could be placed in the same categories based on their

This study investigated the effect of puka gum (PG) (crude and purified) concentration on its emulsification properties in terms of droplet size distribution, zeta potential, rheological and microstructural properties of the emulsions and their visual phase separation. Different concentrations of PG (1–10% w/w) were used to stabilize 15% w/w soybean oil-in-water emulsions. The effect of constant shear rate and time on the apparent viscosity of PG solutions was also tested. PG stabilised-oil-in-water emulsions exhibited monomodal size distributions with average droplet sizes (d32) below 2 μm at 4% w/w PG. The apparent viscosity of these emulsions was dependent on gum concentration exhibiting a Newtonian behaviour at low PG concentrations (≤4% w/w), with shear-thinning at higher concentrations, becoming slightly more pronounced above 100s−1 and especially at 10% w/w gum. The loss modulus (G”) was higher than the storage modulus (G’) at all concentrations indicating that the liquid-like behaviour dominated at all frequencies. The apparent viscosity of PG solutions, showed an irreversible loss of viscosity after shearing over a period of time, presumably resulting from chain breakage and molecular weight reduction. PG droplets were negatively charge with zeta-potential values in the range of −30 – (−45) mV. Furthermore, PG-stabilised emulsions remained stable against phase separation for at least 30 days. Crude and purified PG exhibited no major differences in terms of their emulsifying properties at 4% w/w suggesting that a purification process may not be required for food formulations. Overall, PG can be considered as a promising natural emulsifier for emulsion-based foods and beverage products.

A water-soluble mucilage extracted from the leaves of Hoheria populnea was chemically and physically studied. Monosaccharide composition and linkages were determined by high performance anion exchange chromatography (HPAEC), gas chromatography-mass spectrometry (GC-MS) and nuclear magnetic resonance (NMR) spectroscopy. Lacebark mucilage was composed of rhamnose, galactose, galacturonic acid and glucuronic acid (2:1:2:1). Proton and 13C NMR spectroscopy, and linkage analysis, revealed a predominantly rhamnogalacturonan I-type (RG I-type) structure comprising of a backbone of →4]-α-D-GalpA-[1→2]-α-L-Rhap-[1→. Data indicated the mucilage likely comprises of a polymer containing several structurally discrete domains or possibly more than one discrete polymer. One domain contains a RG I-type backbone with branching at O-3 of GalpA residues to terminal β-D-GlcpA residues, another similarly contains a RG I-type backbone but is branched at O-4 of the Rhap residues to terminal GalpA residues or oligosaccharides containing α-linked 4-Galp and terminal GalpA residues. A possible third domain contains contiguous 2-Rhap residues, some branched at O-3. Hydrated mucilage exhibited pseudoplastic flow behaviour and viscoelastic properties of an entangled biopolymer network. These rheological behaviours were only slightly affected by pH and may prove advantageous in potential end-product applications including oral pharmaceuticals or as a food ingredient.

We evaluated eight biorefinery processes targeting the extraction of ulvan from Ulva ohnoi. Using a factorial design the effect of three sequential treatments (aqueous extraction of salt; ethanol extraction of pigments; and Na2C2O4 or HCl (0.05 M) extraction of ulvan) were evaluated based on the yield (% dry weight of biomass) and quality (uronic acid, sulfate, protein and ash content, constituent sugar and molecular weight analysis) of ulvan extracted. The aqueous extraction of salt followed by HCl extraction of ulvan gave higher yields (8.2 ± 1.1% w/w) and purity of ulvan than equivalent Na2C2O4 extracts (4.0 ± 1.0% w/w). The total sugar content of HCl extracts (624–670 μg/mg) was higher than Na2C2O4 extracts (365–426 μg/mg) as determined by constituent sugar with ulvan specific monosaccharides contributing 94.7–96.2% and 70.1–84.0%, respectively. Ulvan extracted from U. ohnoi was 53.1 mol% rhamnose, 27.8 mol% glucuronic acid, 10.1 mol% iduronic acid, and 5.3 mol% xylose with molecular weights ranging from 10.5–312 kDa depending on the biorefinery process employed. Therefore, the extraction of high quality ulvan from U. ohnoi is facilitated by an aqueous pre-treatment and subsequent HCl-extraction of ulvan as part of a cascading biorefinery model delivering salt, ulvan, and a protein enriched residual biomass.

Manuka honey (MH), α-cyclodextrin (C) and a formulation containing these two components (MH + C) were subjected to simulated gastrointestinal digestion followed by fermentation with human faecal microbiota. The honey monosaccharides, glucose and fructose were 9- and 3-fold higher respectively in the digesta of MH + C compared with MH. Methylglyoxal (MGO), characteristic of MH was absent after gastric digestion. The precursor of MGO, 1,3-dihydroxyacetone was found to be at a higher concentration in MH + C, compared with MH, after digestion. The MH + C fermenta were more acidic (pH 4.6, p

Summary: This protocol describes the application of breath testing and ex vivo fermentations to study the association between breath methane and the composition and functionality of the gut microbiome. The protocol provides a useful systems biology approach for studying the gut microbiome in humans, which combines standardized methods in human breath testing and fecal sampling. The model described is accessible and easy to repeat, but its relative simplicity means that it can deviate from human physiological conditions.

The depolymerisation of porcine mucosal heparan sulfate under the action of heparin lyases and analysis by size-exclusion chromatography (SEC) is described. Heparan sulfate treated to enzymic bond scission producing a Δ 4,5 double-bond and quantified by SEC with ultraviolet-visible (UV) spectroscopic detection (230 nm) indicated that the majority of the biopolymer (>85%) was reduced to disaccharides (degree of polymerisation (DP) = 2). However, analysis of the SEC eluant using refractive index (RI), which reflects the mass contribution of the oligosaccharides rather than the molar response of a UV chromophore, indicated that a considerable proportion of the digested HS, up to 43%, was present with DP >2. This was supported by a mass balance analysis. These results contradict the accepted literature where “complete digestion” is routinely reported. Herein we report on the composition and methodology utilised to ascertain the extent of depolymerization and disaccharide composition of this important biopolymer.

Puka gum (PG), a high molecular weight arabinogalactan-protein (AGP) polysaccharide extracted from the native New Zealand Meryta sinclarii tree formed electrostatic complexes with whey protein isolate (WPI) under specific conditions. The PG and WPI mixtures were slowly acidified using glucono-delta-lactone (GDL), and the progressive change from soluble polymer solution to soluble complexes and to the formation of insoluble complexes (through the association of soluble complexes) were monitored using visual examination, turbidity, particle size (dynamic light scattering) and zeta-potential measurements at total biopolymer concentration C p = 0.3% w/w, protein:polysaccharide ratio Pr:Ps = 2:1, [GDL] = 0.14% w/w, and T = 20 °C. This process was characterised by identifying the phase boundaries, pH c (pH = 5.7 at which soluble complexes of WP and PG were formed) and pH Φ (pH = 4.7 at which complexes associated to form coacervates). The coacervates formed under these conditions appeared as a sticky, viscoelastic and highly extensible material. The effect of ionic strength (0–100 mM NaCl) on complex formation was also studied and a pH c /pH Φ -ionic strength phase diagram was constructed. Coacervation was fully suppressed at ionic strengths above 60 mM NaCl. Rheological measurements of the insoluble coacervate phase at different final coacervation pH (Pr:Ps = 2:1 and 4:1; C p = 3.0% w/w) showed highest complex modulus G * at approx. pH 3.6 for both Pr:Ps = 2:1 and 4:1, i.e. electrical equivalence pH (EEP), where electrostatic interactions are strongest and the overall charge on the insoluble complex is neutral.

A shear-thickening water-soluble polysaccharide was purified from mucilage extracted from the fronds of the New Zealand black tree fern ( Cyathea medullaris or ‘mamaku’ in Māori) and its structure characterised. Constituent sugar analysis by three complementary methods, combined with linkage analysis (of carboxyl reduced samples) and 1 H and 13 C nuclear magnetic resonance spectroscopy (NMR) revealed a glucuronomannan comprising a backbone of 4-linked methylesterified glucopyranosyl uronic acid and 2-linked mannopyranosyl residues, branched at O-3 of 45% and at both O-3 and O-4 of 53% of the mannopyranosyl residues with side chains likely comprising terminal xylopyranosyl, terminal galactopyranosyl, non-methylesterified terminal glucopyranosyl uronic acid and 3-linked glucopyranosyl uronic acid residues. The weight-average molecular weight of the purified polysaccharide was ∼1.9 × 10 6 Da as determined by size-exclusion chromatography coupled with multi-angle laser light scattering (SEC–MALLS). The distinctive rheological properties of this polysaccharide are discussed in relation to its structure.

To date, the structures of the glycolipids from Lactobacillus plantarum, a commonly used beneficial probiotic, have not been conclusively assigned. Herein, we report for the first time, the full characterisation of the four principal glycolipids of the L. plantarum cell wall using sugar, linkage and FAME analysis, as well as ESI-MS/MS and 1D- and 2D-NMR spectroscopy, and assign the major glycolipids as being: α-D-Glcp-diglyceride, α-D-Galp-(1→2)-α-D-Glcp-diglyceride, β-D-Glcp-(1→6)-α-D-Galp-(1→2)-6-O-acyl-α-D-Glcp-diglyceride and β-D-Glcp-(1→6)-α-D-Galp-(1→2)-α-D-Glcp-diglyceride.

Cell wall polysaccharides are resistant to digestion and absorption in the human small intestine and are considered to be delivered to the colon in a chemically unaltered state. In this paper, pulp from green and gold kiwifruit was subjected to in vitro upper-intestinal tract digestion and the chemical and physical changes to cell wall polysaccharides (dietary fibre) were investigated. Yields of insoluble fibre decreased slightly with simulated digestion while soluble fibre yields increased. Constituent sugar and glycosyl linkage analysis of the soluble and insoluble fibre fractions revealed that the chemical composition and structure of the non-starch polysaccharides remained largely unchanged. However, the degree of methylesterification of galacturonic acid residues present in the pectin-rich soluble fibre fractions of both fruit decreased with treatment; size-exclusion chromatography detected changes in the molecular weight profiles of these fractions. These changes may affect the physicochemical properties and fermentability of kiwifruit dietary fibre in the large intestine.

Fractionation of manuka, kanuka and clover honeys indicated the >10 kDa fraction contained small amounts of type II arabinogalactans (AGs), which are often present as arabinogalactan proteins (AGPs). AGPs were isolated from the >10 kDa fraction of kanuka honey using β-glucosyl Yariv reagent and their composition and structure analysed. Constituent sugar, glycosyl linkage and NMR spectroscopy analysis of the purified AGP fraction revealed a predominance of neutral sugars, mainly galactose and arabinose, linked in a highly-branched structure typical of type II AGs. The molecular weight of the major component of the purified AGPs was ∼110 kDa, as determined by size-exclusion chromatography-multi-angle laser light scattering (SEC-MALLS). The Yariv supernatant fraction contained less total sugar, especially galactose, and more protein than purified AGPs. Linkage analysis indicated this fraction also contained an AG-type polymer in addition to various other polysaccharides and SEC-MALLS indicated the molecular weight of the major component was ∼32 kDa.

The cell-wall polysaccharide components of gold kiwifruit ( Actinidia chinensis Planch. ‘Hort16A’, ZESPRI ® GOLD) have been investigated for the first time. Alcohol-insoluble residues (AIRs) were prepared from whole, unpeeled gold and green ( Actinidia deliciosa ) kiwifruit and the constituent sugar and glycosyl linkage compositions determined. AIRs from both kiwifruit contained a high proportion of cellulose; the gold kiwifruit contained a higher proportion of hemicellulosic polysaccharides and lower proportion of pectic polysaccharides compared with the green. The gold kiwifruit AIR was partitioned by sequential extraction with water, aqueous CDTA (0.05 M), aqueous Na 2 CO 3 (0.05 M) and aqueous KOH (1 M then 4 M). The glycosyl linkage compositions of each fraction were determined and the types of cell-wall polysaccharides from the gold fruit were found to be similar to those previously reported for the green fruit.

The water-soluble carbohydrates (WSCs) extracted from the underground parts (rhizome) of Arthropodium cirratum (Rengarenga lily extract); third order branches of Cordyline australis (Cabbage tree extract); a seaweed, Undaria pinnatifida (Undaria extract), and exudates from Acacia pycnantha (Acacia extract) were investigated. Extracts of Rengarenga lily, Cabbage tree, Undaria, and Acacia contained 576, 250, 275 and 794 g/kg DM WSCs, respectively. Constituent sugar analysis by gas–liquid chromatography (GLC) showed that extracts of Rengarenga lily and Cabbage tree contained predominantly fructose and glucose (82–95%). The analysis also revealed that Acacia extract contained mainly galactose (78%) and arabinose (22%) while Undaria extract, contained fucose (55%) and galactose (44%). Thin-layer chromatography (TLC) showed that, on the basis of RF values, fructan composition of Rengarenga lily extract and Cabbage tree extract was different. Cabbage tree extract contained 45% (w/w) fructans while Rengarenga lily extract contained 65% (w/w) fructans. High performance size-exclusion chromatography coupled with multi-angle laser light scattering (SEC-MALLS) showed that the extracts had varying weight average molecular weight due to differences in the average chain length of the major carbohydrates. Data for the amino acid compositions differed considerably depending on the type of extract. Water-soluble carbohydrate extracts prepared from the four plant sources gave a wide range of WSC (250–794 g/kg DM) due to the different proportions of structural material in different species. It is not known how these differences will impact on animal production, if diets are supplemented with the extracts.

The polysaccharides exuded from both Phormium tenax and Phormium cookianum leaves are acidic xylans with molecular weights in the range 1.3×10 6 –1.7×10 6 Da. 1 H and 13 C NMR spectra show that in both cases the backbone comprises 1,4-linked β- d -Xyl p units and their substituted products, including a structural unit that is both 2- O -substituted and 3- O -acetylated. The sidechains contain α- d -GlcA and are terminated by β- d -Xyl p and α- l -Ara f units in relative proportions that differ between plant varieties. The polysaccharides are so highly branched that chain-terminating units account for approximately half of all structural units. These properties place the Phormium polysaccharides in a class which includes brea, sapote and yabo gums.

Casein-maltodextrin glyco-conjugates were prepared using an economical, food-grade process based on the Amadori re-arrangement of the Maillard reaction. The resultant glyco-conjugates were slightly yellow in colour and the degree of discolouration was dependent on heating time. Formation of glyco-conjugates was demonstrated by determining the reduction of free amino-groups by the O-phthaldialdehyde (OPA) assay and sugar reducing ends by gas chromatography-mass spectrometry (GC-MS). Increases in molecular weight were monitored by SDS-polyacrylamide gel electrophoresis (PAGE) and were in agreement with those predicted for the conjugation of casein monomers with malto-oligosaccharides of average DP 7-10. 2D-Urea-SDS-PAGE demonstrated that both protein and saccharide components co-migrate, indicating that covalent bonds were upon heating. This resulted in increases in mass-to-charge ratio of the materials, which suggested decreases in pI. These observed chemical and physical changes were reconciled with previously documented improvements in emulsifying properties. © 2004 Elsevier Ltd. All rights reserved.

A gum that exudes from the wounded trunk of the New Zealand native tree Meryta sinclairii has been isolated. The gum was completely precipitated by the β-glucosyl Yariv reagent and was thus determined to be an arabinogalactan-protein (AGP). It contained >95% w/w carbohydrate and only 2% w/w protein with a high level of hydroxyproline. SEC-MALLS showed that the gum had a weight-average molecular weight of 4.45×10 6 Da compared with 6.02×10 5 Da for gum arabic. Constituent sugar and linkage analyses were consistent with polymers comprised of a highly branched backbone of 1,3-linked galactopyranosyl (Gal p ) residues, with side-chains made up of arabinofuranose- (Ara f ) containing oligosaccharides, terminated variously by rhamnopyranosyl (Rha p ), arabinopyranosyl (Ara p ), Gal p and glucuronopyranosyl (Glc p A) residues. Analysis by one-dimensional and two-dimensional 1 H and 13 C NMR experiments confirmed the linkage analyses. The structure of the gum is discussed in comparison with the structure of gum arabic and other AGPs.

At about 20-year intervals, since at least the 1860s, there have been accounts of the accumulation of large quantities of mucilage in the waters off the northern coast of the South Island of New Zealand. On a few occasions these events have been associated with harmful effects, such as the mass mortality of marine fauna and the impediment of fishing activities, though previous attempts to identify the causative organism(s) or the environmental conditions which promote its development have been unsuccessful. In January 2000, there was a moderate re-occurrence of mucilage accumulation in Tasman Bay that led to the identification of the primary origin of this material as the planktonic, thecate, dinoflagellate Gonyaulax hyalina (Ostenfeld et Schmidt). Macro-aggregate accumulation was occurring in the water column when dinoflagellate cell numbers were relatively low. Experimental work on cultures of this organism showed that the dinoflagellate produced transparent exo-polymers (TEPs) composed primarily of galactose and glucose monomers (44.8 and 29.6 mol%, respectively). Analysis of natural and cultured mucilage showed this had a high inorganic content (>60% (w/w)). A substantial proportion (40% (w/w)) of this inorganic fraction was composed of calcium that apparently played an important role in stabilizing the gel. Phosphorus was also enriched in the mucilage. It is hypothesized that the high inorganic N/P ratio observed in the water column at the time of mucilage formation was an effect rather than a cause of this event, which resulted in the scavenging of particulate material from the water column and enrichment of sediments and bottom waters with organic material and phosphorus.

Summary Starch granules isolated from leaves and seeds of Lolium temulentum L. were roughly spherical with an irregular perimeter having mean diameters of 1.7 and 4.1 μm respectively. On a dry mass basis, 47 % of the seed and maximally 4 % of the leaf were starch. Starch structures were deduced from a combination of data obtained using acid-hydrolysis/Smith degradation, glycosyl-linkage analysis/GC-MS, iodine complex formation, isoamylase digestion, high performance size exclusion chromatography and high performance anion exchange chromatography/pulsed amperometry. The mean degree of polymerisation (DP) of seed starch was 4800 (Mr = 778 kDa) and consisted entirely of a highly branched, amylopectin-like polymer with an average chain length of DP = 46 ± 6 and an average of 104 chains per molecule. Leaf starch was 98.8 % by mass of a highly-branched, amylopectin-like polymer of DP = 5850 (Mr 947 kDa) containing on average, 136 chains of mean DP = 43 ± 6. The remaining 1.2 % of the leaf starch was isoamylase-resistant, of high Mr (5850 kDa) and was designated amylose. Both starches were unusual in that the chain lengths were longer by a factor of 2, than is normal for amylopectins and also because they contained little or no long 1,4-linked chains, as determined by isoamylase digestion. HPAEC of isoamylase fragments from leaf starch revealed a bi-modal distribution of branch chain lengths which was not evident in the seed starch. With the exceptions of this bi-modality and the trace of amylose in the leaf starch, the structure of the starches from the two organs were very similar. Seed starch exhibited an anomalous reaction with iodine. The chromogen was deep blue with a λmax = 608 nm, indicative of the presence of long amylose chains. On digestion with isoamylase, all of the mass of starch was converted to low molecular mass with an iodine λmax shifted to 567 nm. Hence, short 1,4-linked glucan chains which were contiguous through 1,6-linkages formed blue chromogens and can give the appearance of amylose.

Fermentation of Gluconobacter oxydans NCIMB 4943 using malto-oligosaccharides of differing average degrees of polymerisation (DP) as substrates resulted in the synthesis of dextrans consistent with the action of the enzyme dextran–dextrinase (DDase). These dextrans could be precipitated from the fermentation broth using Fehling's solution. The glycosyl linkage composition of the dextrans was largely independent of the size of the initial substrate used and fermentation time. They contained an average of 83 mol% 6-linked, 8 mol% 4-linked, 6 mol% terminal and 3 mol% 4,6-linked glucopyranosyl residues. The dextrans gave viscous solutions that displayed shear-thinning behaviour. In addition to the synthesis of dextrans, the fermentations of G. oxydans with malto-oligosaccharides also resulted in the formation of oligosaccharides that were soluble in Fehling's solution. These oligosaccharides contained varying proportions of 4-Glc p and 6-Glc p residues depending on the size of the initial substrate and the culture time. These oligosaccharides may have prebiotic properties.

The polymers secreted by suspension-cultured apple cells were composed of 85% carbohydrate (76% neutral sugar and 9% uronic acid) and 15% w/w protein. The extracellular polysaccharides (ECPs) contain 23% XG and 59% AGPs. The monosaccharide composition of the ECPs consisted of Gal, Ara, Glc and Xyl, with smaller amounts of Rha, Fuc and Man. Fractionation of the ECPs by anion-exchange chromatography yielded an unbound neutral fraction and a bound acidic fraction. Monosaccharide and linkage compositions of each fraction were determined. The neutral fraction (48% recovered carbohydrate) was composed of xyloglucan (XG; >90 mol%) which was purified by selective precipitation with Fehling’s solution to yield pure XG. The purified XG had a Glc:Xyl:Gal:Fuc ratio of 4.0:2.5:0.8:0.5; the XG was not O- acetylated. The structure of the secreted XG was similar to that extracted from apple-pomace. The acidic fraction (52% recovered carbohydrate) was composed primarily of arabinogalactan-proteins (AGPs) as detected by the β -glucosyl Yariv diffusion test. The AGP had a Gal:Ara ratio of 1.3: 1.0. Minor amounts of arabinan, xylan and mannan were also detected in the ECPs. This study is the first examination of the polysaccharides secreted by apple cells grown in suspension culture.

The rheological properties of three xyloglucans (XGs) from the extracellular medium of suspension cultured Nicotiana plumbaginifolia cells, apple pomace and tamarind seeds, with different structural features and molecular weights have been studied. The molecular weight (weight average) of the Nicotiana , apple pomace and tamarind seed XGs determined by multi-angle laser light scattering were 129, 219 and 833 kDa, respectively. Tamarind seed XG had the highest viscosity and Nicotiana XG had the lowest viscosity, with that of apple pomace XG intermediate. The viscosity of apple pomace XG at 5% w/v was almost equivalent to that of tamarind seed XG at 2% w/v, but their behaviour at high shear rates differed; both XGs were non-Newtonian in their rheological properties, but that from tamarind seeds showed more pronounced shear-thinning. The viscosity of Nicotiana XG at 5% w/v was almost equivalent to tamarind seed XG at 0.5% w/v, displaying Newtonian behaviour. Modification of the molecular weight of the XGs and their degree of branching revealed that differences in viscosity between the molecules, and their shear-field behaviour, was due primarily to differences in molecular weight. Removal of fucose residues from apple pomace XG decreased the viscosity of solutions from 8 to 4 mPa·s, whereas removal of both fucose and galactose from apple pomace XG, resulted in precipitation from solution. Deacetylation of Nicotiana XG also resulted in precipitation from solution.

The soluble polymers secreted by cell-suspension cultures of Nicotiana plumbaginifolia contained 78% carbohydrate, 6% protein and 4% inorganic material. The extracellular polysaccharides were separated into three fractions by anion-exchange chromatography using a gradient of imidazole-HCl at pH 7 and the individual polysaccharides in each fraction were then isolated by selective precipitation and enzymic treatment. Monosaccharide and linkage compositions were determined for each polysaccharide after reduction of uronic acid residues and the degree of esterification of the various uronic acid residues in each polysaccharide was determined concurrently with the linkage types. Six components were identified: an arabinoxyloglucan (comprising 34% of the total polysaccharide) and a galactoglucomannan (15%) in the unbound neutral fraction, a type II arabinogalactan (an arabinogalactan-protein, 11%) and an acidic xylan (3%) in the first bound fraction, and an arabinoglucuronomannan (11%) and a galacturonan (26%) in the second bound fraction.

Individual fructan tri-, tetra- and pentasaccharide isomers in neutral, water-soluble extracts from Lolium temulentum were purified and the linkages present in these isomeric oligosaccharides were analysed by combined GC-mass spectrometry of partially methylated alditol acetates. 1-Kestose and neokestose were the most abundant trisaccharides with 6-kestose present in much lower amounts. Analysis of isomers of DP 4 and 5 showed that multiple linkage types were present with structures based on all three trisaccharides. Oligosaccharides based on neokestose but with 2,6 linkages between adjacent fructose residues have not been previously detected in higher plants.