Structural studies on the specific capsular polysaccharide from Rhizobium trifolii, TA-1

The repeating unit of the specific capsular polysaccharide from the bacterium Rhizobium trifolii (TA)-1 has been shown to contain (a) terminal 4,6-O-(1-carboxyethylidene)-D-galactose (1 residue), (b) (1 → 3)-linked 4,6-O-(1-carboxyethylidene)-D-glucose (1 residue), (c) (1 → 4)-(1 → 6)-linked D-glucose (1 residue), (d) (1 → 4)-linked D-glucuronic acid (1 residue), and (e) (1 → 4)-linked D-glucose (4 residues). The pyruvylated sugars were shown to be positioned sequentially, and at least one other unit was interposed between them and the branch point.     

Mammalian-lung galactan

Exopolysaccharides of Agrobacterium tumefaciens and Rhizobium meliloti, containing d-glucose, d-galactose, pyruvic acid, and O-acetyl groups in the approximate proportions 6:1:1:1.5, were analysed by methylation. They were found to contain the following main structural units (all β-glycosidic): chain residues of (1→3)-linked d-glucose (24%), (1→3)-linked d-galactose (15%), (1→4)-linked d-glucose (20%), and (1→6)-linked d-glucose (18%); (1→4,1→6)-linked branching residues of d-glucose (12%), and terminal d-glucose residues substituted at positions 4 and 6 by pyruvate (11%). Uronic acid-containing exopolysaccharides of Rhizobium leguminosarum, R. phaseoli, and R. trifolii contained d-glucose, d-glucuronic acid, d-galactose, pyruvic acid, and O-acetyl groups in the approximate proportions 5:2:1:2:3. Methylation gave identical patterns of methylated sugar components, from which the following structural elements were deduced: chain residues of (1→3)-linked d-glucose substituted at positions 4 and 6 by pyruvate (13%), (1→4)-linked d-glucose (32%), and (1→4)-linked d-glucuronic acid (20%); (1→4,1→6)-linked branching residues of d-galactose and/or d-glucose (13%), and terminal d-glucose and/or d-galactose residues substituted at positions 4 and 6 by pyruvate (13%).     

Some structural features of a new sulphated heteropolysaccharide from Padina pavonia

An acid-extractable, water-soluble, polysaccharide sulphate, isolated from Padina pavonia, comprised variable proportions of glucuronic acid, galactose, glucose, mannose, xylose, and fucose in addition to a protein moiety. Partial acid hydrolysis and autohydrolysis of the free acid polysaccharide yielded several oligosaccharides. Evidence from periodate oxidation studies indicated that the inner polysaccharide portion is composed of (1 → 4)-linked β-D-glucuronic acid, (1 → 4)-linked β-D-mannose and (1 → 4)-linked β-D-glucose residues. The heteropolymeric partially sulphated exterior portion is attached to the inner part and comprises various ratios of (1 → 4)-linked β-D-galactose, β-D-galactose-3-sulphate residues, (1 → 4)-linked β-D-glucose residues, (1 → 2)-linked α-L-fucose 4-sulphate residues and (1 → 3)-linked β-D-xylose residues.     

The molecular structures of a glucan and a galactan synthesised by Prototheca zopfii

When the unicellular organism Prototheca zopfii was grown on a malt-agar medium, a mixture of polysaccharides was synthesised which could be subsequently extracted from the dried cells with hot water and hot alkali. The major polysaccharide was a galactan which had a branched structure with main chains of (1→6)-linked D-galactopyranose residues, and ≈ 10% of side chains containing terminal D-galacto-furanose residues. A glycogen-type polysaccharide and a (1→4)-linked mannan were also produced.     

Structural investigations of the extracellular polysaccharide elaborated by s19, a Xanthomonas-type bacterium

The extracellular, acidic heteropolysaccharide from Xanthomonas S19 consists of D-glucuronic acid, D-glucose, D-galactose, and D-mannose residues in the approximate molar ratios of 1.6:3:1:1, plus acetyl groups liked to C-2 and/or C-3 of a large proportion of the glucose residues. Methylation studies showed that the glucose is present as non-reducing end-group also as 1,2- and 1,4-linked units, the galactose residues are solely 1,3-linked, a major proportion of the mannose residues are 1,2,4-linked and the rest 1,2-linked. A high proportion of the glucuronic acid units are 1,4-linked. Periodate oxidation confirmed the presence of these linkages. The disaccharides D-Glc-(1→4)-D-Glc,D-Glc-(1→2)-D-Man, D-Glc-(1→3)-D-Gal, D-Gal-(1→2)-D-Glc, D-GlcA-(1→4)-D-GlcA, and β-D-GlcA-(1→4)-D-Man were isolated from a partial hydrolysate of the polysaccharide, and characterised. The similarities and differences between this polysaccharide and those from other Xanthomonas species are discussed.     

Smith degradation of gum exudates from some prosopis species

The polysaccharide of P. hymantophora has been shown to be composed of (1→4)-linked galactopyranosyl, (1→3)-linked galactopyranosyl, (1→3)-linked galactopyranosyl 2- and 4-sulphate and 2,6-disulphate residues. The (1→3)- and (1→4)-linked units are present in approximately equal amounts. The polysaccharide of P. hieroglyphica has been shown to possess (1→4)-linked galactopyranosyl, (1→3)-linked galactopyranosyl, and (1→3)-linked galactopyranosyl 2- and 4-sulphate residues. The (1→3)- and (1→4)-linked units are present in a 4:1 ratio. Both polysaccharides contain small proportions of non-reducing xylosyl end-groups.     

An arabinogalactan from the seeds of Pseudium guava

Hemicelluloses of seeds of Pseudium guava containing d-galactose (59.6), d-arabinose (35.9), and a uronic acid (4.5%) were analyzed by methylation analysis and Smith-degradation analysis, and the following structural elements were deduced; chain residues of (1→4)-linked d-galactose, (1→5)-linked d-arabinose, and terminal d-arabinose residues. The following structure was assigned to the polysaccharide. →5)-d-Araf-(1→4)-d-Galf-(1→5)-d-Araf-(1→     

Analytical and structural features of the gum exudate from Combretum hartmannianum schweinf.

The gum exudate from Combretum hartmannianum is water-soluble, forms very viscous solutions, and contains galactose (22%), arabinose (43%), mannose (10%), xylose (6%), rhamnose (4%), glucuronic acid (6%), 4-O-methylglucuronic acid (2%), and galacturonic acid (7%). The acidic components produced on hydrolysis of the gum were 6-O-(β-D-glucopyranosyluronic acid)-D-galactose, and two saccharides that had the same chromatographic mobility, and contained mannose and galacturonic acid, and galactose and 4-O-methylglucuronic acid, respectively. Methylation and methanolysis of the gum indicated the presence of terminal uronic acid, rhamnose, xylose, galactose, arabinofuranose, and arabinopyranose. Controlled, acid hydrolysis indicated the presence of (1→3)-linked arabinopyranose side-chains and (1→6)-linked galactose residues. C. hartmannianum gum, when subjected to two Smith-degradations, yielded Polysaccharides I and II, both of which contained galactose, arabinose, and mannose. Insufficient crude gum was available for a complete structural study, but the molecule was shown to contain long, sparsely branched chains of (1→6)-linked galactose residues, to which are attached (1→3)-linked arabinose and (1→3)-linked mannose side-chains.     

Structural investigations on the lignin–carbohydrate complexes of Lolium perenne

1. Lignin-carbohydrate complexes isolated from leaf blade, leaf sheath and stem tissue of ryegrass by extraction with dimethyl sulphoxide were examined by fractionation procedures. Although the complexes are heterogeneous, heterogeneity is shown only in the ratio of the individual monosaccharide residues and not in the ratio of lignin to carbohydrate. 2. The molecular weight of the complexes is high (>/=150000), but chemical modification by alkaline hydrolysis, borohydride reduction or lead tetra-acetate oxidation does not drastically decrease it. Low-molecular-weight fragments released by alkaline treatment were shown to contain acetic acid, ferulic acid and p-coumaric acid. 3. On the basis of the chemical stability of the complexes, it is postulated that at least three types of bonding may be present between lignin and carbohydrate, namely one cleaved on borohydride reduction, another cleaved by alkali and a linkage resistant to alkali. 4. The carbohydrate portion of the complexes is composed of beta-(1-->4)-linked d-glucose residues (cellulose) and beta-(1-->4)-linked chains of xylose residues. Side chains involving arabinose and galactose residues are linked to C-3 of some of the xylose residues. 5. How the components of the complexes are held together is not certain, but it is suggested that the phenolic acids may act as cross-linking agents.     

Structural features of two amyloids from the hemi-cellulosic fraction of field-bean (Dolichos lablab) hulls

Two amyloid-type fractions were isolated from field-bean (Dolichos lablab) hulls by 10% alkali extraction followed by acetylation and solvent fractionation. The major, chloroform-insoluble fraction and a minor, chloroform-soluble fraction were found to be homogeneous in sedimentation analysis and molecular-sieve chromatography. The polysaccharides contained xylose and glucose in various proportions. Methylation analysis, periodate oxidation, Smith degradation, oxidation by chromium trioxide, and oligosaccharide studies indicated a new type of structure for the major fraction (glucose:xylose ratio of 1.9:1) in that it had a backbone of (1→4)-linked β-d-glucose residues interspersed with single or multiple residues of (1→4)-linked β-d-xylose, and to which some single d-xylosyl groups are attached through O-6 of d-glucose. In contrast, the minor fraction (glucose:xylose ratio of 1:3.7) had a backbone of (1→4)-linked β-d-xylose interspersed with (1→4)-β-d-glucose and having a side chain of d-xylose, attached through O-6 of d-glucose. The third fraction was found to be a mixture of linear (1→4)-d-glucan and (1→4)-d-xylan.     

Characterisation of the extractable pectins and hemicelluloses of the cell wall of carrot

Pectic and hemicellulosic polysaccharides were successively extracted from an alcohol-insoluble residue (AIR) from carrot root by the actions of Pronase, hot dilute acid, cold dilute alkali, and concentrated alkali in yields corresponding to 12.6, 13.5, 21.7, and 6.7% of AIR, respectively. The first two products were fractionated further by ion-exchange chromatography. Carrot pectins contained 61.3–66.0% of galacturonic acid and 16.0–19.9% of neutral sugars, mainly galactose, arabinose, and rhamnose. Except for the alkali-soluble pectins, the degrees of methylation were high (62.9–67.1) and there was a significant degree of acetylation (7.2–13.5). Pectin fractions were homogeneous in gel-filtration chromatography with viscosity-average molecular weights varying between 36,200 and 56,500. Methylation analysis indicated the presence of arabinogalactans in the pectins extracted during the proteolysis, and fairly long chains of (1→4)-linked galactan with a branched arabinan in the two other pectic fractions. The hemicellulose fraction was mainly composed of (1→4)-linked glucan, (1→4)-linked mannan, (1→4)-linked xylan, and small but significant amounts of pectic polysaccharides. The possible association of cell-wall polymers is discussed.     

Constitution of sargassan,a sulphated heteropolysaccharide from sargassum linifolium

The behaviour towards periodate of the brown-algal polysaccharide sargassan before and after partial hydrolysis, alkali treatment, and methanolysis has been studied. Evidence is thereby provided that the sargassan backbone is composed of (1→4)-linked β-D-glucuronic acid and β-D-mannose residues. Heteropolymeric, partially sulphated branches are attached to the backbone, and these branches comprise various proportions of(l→4)-linked, β-D-galactose, β-D-galactose 6-sulphate, and β-D-galactose 3,6-disulphate residues, (1→2)-linked α-L-fucose 4-sulphate residues, and (1→3)-linked β-D-xylose residues.     

Structural features of a sulphated,fucose-containing polysaccharide from the brown seaweed dictyota dichotoma

A sulphated heteropolysaccharide (～15% of the acid-extractable material) isolated from the brown alga Dictyota dichotoma contains residues of D-glucuronic acid, D-galactose, D-mannose, D-xylose, and L-fucose¹. Partial hydrolysis of the polysaccharide with acid gave one neutral and two acidic oligosaccharides. The behaviour towards periodate of the polysaccharide before and after partial hydrolysis, alkali-treatment, and methanolysis has been studied. Evidence is thereby provided that the polysaccharide is partially sulphated and composed of (1→4)-linked residues of D-glucuronic acid, D-galactose, D-mannose, and D-xylose, and (1→2)-linked L-fucose.     

Chemical structures in a lignin-carbohydrate complex isolated from the bovine rumen

The soluble, lignin-carbohydrate complex (LCC) from the rumen fluid of steers fed a diet of pure spear grass (Heteropogon contortus) has been purified by gel filtration. The purified LCC contained 7.4% of carbohydrate which, on hydrolysis, gave d-glucose, d-xylose, l-arabinose, l-rhamnose, and traces of d-galactose and d-mannose. The structure of the LCC was examined by methylation analysis, using g.l.c.-m.s. for the unequivocal classification of the sugar derivatives. d-Glucose, d-xylose, and l-rhamnose were shown to be glycosidically linked to lignin. Some of the d-glucosyl residues carry other (1→4)-linked d-glucose units, and some of the d-xylosyl residues bear other (1→4)-linked d-xylose units and (1→3)-linked l-arabinofuranosyl groups. The major carbohydrate component is a single d-glucopyranosyl group. The LCC was subjected to various chemical treatments in an investigation of the chemical nature of the bonding between lignin and the carbohydrates. d-Glucose could be enzymically hydrolyzed from the LCC, but only with a very high concentration of β-d-glucosidase. The presence of lignin in rumen LCC has been confirmed by nitrobenzene oxidation, vanillin and syringaldehyde being identified by g.l.c.-m.s. as oxidation products from both the original spear grass and the LCC.     

Structures and Antitumor Activities of the Polysaccharides Isolated from Fruiting Body and the Growing Culture of Mycelium of Ganoderma lucidum

Several β-D-glucans, appertaining to the same molecular species but having different degrees of branching, were isolated from water and alkali extracts of the fruiting body of Ganoderma lucidum (Reishi). The purified glucans that were mostly water-insoluble had a backbone of (1 →3)-linked D-glucose residues, attached mainly with single D-glucosyl units at 0-6 and also with a few short (l→4)-linked glucosyl units at 0-2 positions. However, their degrees of branching appeared to differ in the range of d.b. 1/3 ~ 1/23, depending on the extracted glucan fractions. In addition to the ^-glucans, the fruiting body contained water-soluble heteropolysaccharides, comprising D-glucose, D-galactose, D-mannose, L-(or D)-arabinose, D-xylose, and L-fucose.

A branched (1 →3)-β-D-glucan was also isolated from the culture filtrate of G. lucidum grown in a glucose-yeast extract medium. The extracellular β-D-glucan was less soluble in water after purification, but soluble in dilute alkali. This glucan has essentially the same structure as that of hot-water extracted polysaccharide from the fruiting body. The repeating unit of the glucan contains a backbone chain of (1 →3)-linked D-glucose residues, five out of sixteen D-glucose residues being substituted at 0-6 positions with single D-glucosyl units and one D-glucose residue at 0-2 positions probably with a cellobiose unit.

The hot-water extractable fruiting body glucan and the extracellular glucan of the culture of growing mycelium showed relatively high growth-inhibition activities against Sarcoma 180 solid tumor in mice, when administered by. successive intraperitoneal injections. When the moderately branched glucans were modified to D-glucan-polyols by periodate oxidation and borohydride reduction, they exhibited higher antitumor activities, confirming the previous conclusion that the attachment of polyol groups to the (1 →3)-lmked backbone significantly enhances its host-mediated antitumor effect.     

Structure of l-arabino-d-galactan-containing glycoproteins from radish leaves

Two l-arabino-d-galactan-containing glycoproteins having a potent inhibitory activity against eel anti-H agglutinin were isolated from the hot saline extracts of mature radish leaves and characterized to have a similar monosaccharide composition that consists of l-arabinose, d-galactose, l-fucose, 4-O-methyl-d-glucuronic acid, and d-glucuronic acid residues. The chemical structure features of the carbohydrate components were investigated by carboxyl group reduction, methylation, periodate oxidation, partial acid hydrolysis, and digestion with exo- and endo-glycosidases, which indicated a backbone chain of (1→3)-linked β-d-galactosyl residues, to which side chains consisting of α-(1→6)-linked d-galactosyl residues were attached. The α-l-arabinofuranosyl residues were attached as single nonreducing groups and as O-2- or O-3-linked residues to O-3 of the β-d-galactosyl residues of the side chains. Single α-l-fucopyranosyl end groups were linked to O-2 of the l-arabinofuranosyl residues, and the 4-O-methyl-β-d-glucopyranosyluronic acid end groups were linked to d-galactosyl residues. The O-α-l-fucopyranosyl-(1→2)-α-l-arabinofuranosyl end-groups were shown to be responsible for the serological, H-like activity of the l-arabino-d-galactan glycoproteins. Reductive alkaline degradation of the glycoconjugates showed that a large proportion of the polysaccharide chains is conjugated with the polypeptide backbone through a 3-O-d-galactosylserine linkage.     

Fine structure of (1→3),(1→4)-β-d-glucan from Zea shoot cell-walls

The insoluble material that remains after extraction of Zea shoots with cold buffer was treated successively with 3m LiCl and hot water. The polysaccharides solubilized by these treatments were mostly (1→3),(1→4)-β-d-glucans. The β-d-glucan from the hot-water-soluble fraction was hydrolyzed by Bacillus subtilis (1→3),(1→4)-β-d-glucan 4-glucanohydrolase. The oligosaccharides were characterized by methylation analysis of the enzymic fragments and by methylation analysis of secondary fragments generated by treatment of the isolated oligosaccharides with Streptomyces QM B814 cellulase. The results demonstrate that the native polysaccharide consists mainly of cellotriosyl and cellotetraosyl residues joined by single (1→3) linkages. Evidence is presented to show that certain other glucosyl sequences are also present in the native polysaccharide including (a) two, three, or four contiguous (1→3)-linkages; (b) blocks of more than four (1→4)-linked glucose residues; (c) regions having alternating (1→3)- and (1→4)-linkages.     

Carbohydrates of the antarctic brown seaweed Ascoseira mirabilis

Mannitol, sucrose and four monosaccharides were obtained from an ethanolic extract of Ascoseira mirabilis. Sequential extraction with aqueous calcium chloride, dilute acid and dilute alkali gave mixtures of laminaran, ‘fucan’ and alginic acid. Laminarans fractionated from the extracts contained different proportions of uniformly (1 → 3) and (1 → 6) linked chains of β-D-glucose residues. The ‘fucan’ contained varying proportions of fucose, galactose and glucuronic acid, small amounts of xylose, mannose, glucose, half ester sulphate and protein. Extraction of the weed under mild alkaline conditions gave a yield of 13.4% of low molecular weight calcium alginate with a mannuronate to guluronate ratio of 30:70 and only a small proportion of sequences of alternating residues. Selective extraction and fractionation gave alginate fractions rich (> 80%) in mannuronate or guluronate.     

Treatment of rhamnogalacturonan I with lithium in ethylenediamine

Rhamnogalacturonan I is a pectic polysaccharide that is solubilized from the walls of suspension-cultured sycamore cells (Acer pseudoplatanus) by the action of a highly purified endo-1,4-α-polygalacturonanase. Rhamnogalacturonan I has a linear backbone consisting of the diglycosyl repeating unit, →4)-α-d-GalpA-(1→2)-α-l-Rhap-(1→. Approximately half of the α-l-rhamnosyl residues of the backbone are branched at O-4. Selective cleavage at the galactosyluronic acid residues of the backbone by treatment of rhamnogalacturonan I wit lithium in ethylenediamine resulted in the release of the neutral glycosyl-residue sidechains that had been attached to the backbone. Various analytical techniques, including combined liquid chromatography-mass spectrometry, combined gas-liquid chromatography-mass spectrometry, and ¹H-nuclear magnetic resonance spectroscopy, were used to determine the structure of the side chains. The majority of the sidechains were isolated as oligoglycosylalditols, with rhamnitol at the “reducing” end. Terminal 2-, 4-, or 6-linked galactosyl residues were found attached to O-4 of the rhamnitol residues The 2-, 4-, and 6-linked galactosyl residues had terminal or 2-linked arabinosyl, or additional galactosyl, residues attached to them. Based on the results of fast-atom-bombardment mass spectrometry, the side chains were found to range in size from one to fourteen glycosyl residues. The side-chain structures suggest that there are four or more distinct families of side chains attached to the backbone of rhamnogalacturonan I.     

Characterization of anti-complementary acidic heteroglycans from the seed of Coix lacryma-jobi var. ma-yuen

The anti-complementary polysaccharides, CA-1 and CA-2, were purified from the seed of Coix lacryma-jobi L. var. ma-yuen. CA-1 consists of rhamnose, arabinose, xylose, galactose, galacturonic acid and glucuronic acid in molar ratios of 1.8:43.8:10.8:33.2:3.2:7.2, and CA-2 consists of rhamnose, arabinose, xylose, mannose, galactose, glucose, galacturonic acid and glucuronic acid in molar ratios of 2.4:37.0:11.8:1.7:35.6:2.9:2.6:6.0. CA-1 and CA-2 contained 8 ∼ 11% protein. Their M_rs were estimated to be 160 000 in CA-1 and 70 000 in CA-2 by gel filtration. CA-2 showed more potent anti-complementary activity than CA-1 in low dose.Methylation analysis of CA-1, its carboxyl-reduced products (reduced CA-1a and CA-1b) and CA-2 were carried out by the use of GC/MS and the results suggested that CA-1 has a very complicated and highly branched structure, and CA-2 is also composed of the same glycosidic linkages as CA-1 in different molar proportions. The results of exo α-L-arabinofuranosidase treatment and partial acid hydrolysis suggested that CA-1 and CA-2 contained arabino 3,6-galactan moiety and most of the arabinose was present as an α-L-furanosyl residues in the non-reducing terminals and (1 → 5)-linked side chains which mostly attached to the O-3 of (1 → 6)-linked galactopyranosyl residues. The results also suggested that CA-1 and CA-2 contained rhamnogalacturonan moiety which has a main chain consisting of (1 → 4)-linked galacturonic acid and (1 → 2)-linked rhamnose, and arabino 3,6- and 4-galactan might be attached to the rhamnosyl residue at the O-4. All glucuronic acid residues were present at the non-reducing terminals.