Structural study of the extracellular polysaccharide gum from Rhizobium strain CB744

The structure of the extracellular polysaccharide gum from nitrogen-fixing Rhizobium sp. strain CB744 (a member of the slow-growing Cowpea group) has been investigated. Gas-chromatographic analysis of the alditol acetates of the acid hydrolysate showed the gum to be composed of galactose, 4-O-methylgalactose, mannose, and glucose in the molar ratio of 1:2.5:3.5:7.0. The polysaccharide is unusual in that it contains no carbonyl substituent, although such substituents are common amongst polysaccharides produced by the slow-growing group. The native and de-branched polysaccharides were examined by methylation analysis. The anomeric configurations were determined by ¹³C-n.m.r. and oxidation by chromium trioxide. It is concluded that there are two β-(1→4)-linked glycopyranosyl residues for each α-(1→4)-linked mannopyranosyl residue, and that each mannose is substituted at O-6 by a β-galactopyranosyl residue, with 71% of the galactose groups being present as 4-O-methylgalactose.     

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.     

Pyruvic acid-containing mono- and oligo-saccharides from rhizobium trifolii bart A

After partial, acid hydrolysis of the extracellular, acid polysaccharide from Rh. trifolii Bart A, the following products were isolated and characterized: 3,4-O-(1-carboxyethylidene)-d-galactose, 4,6-O-(1-carboxyethylidene)-d-galactose, 3-O-[3,4-O-(1-carboxyethylidene)-β-d)-galactopyranosyl]-d-glucose, 3-O-[4,6-O-(1-carboxyethylidene)-β-d-galactopyranosyl]-d-glucose, O-[3,4-O-(1-carboxyethylidene)-β-d-galactopyranosyl ]-(1→3)-O-d-glucopyranosyl-(1→4)-d-glucose, and O-[4,6-O-(1- carboxyethylidene)-β-d-galactopyranosyl]-(1→3)-O-β-d-glucopyranosyl-(1→4)-d-glucose. The presence of pyruvic acid linked either to O-3 and O-4 or to O-4 and O-6 of the d-galactopyranosyl group of these saccharides indicates that both structures may be present in the original polysaccharide.     

Structural studies of plant gum from sap of the lac tree, Rhus vernicifera

The plant gum isolated from sap of the lac tree, Rhus vernicifera (China), was separated into two fractions having mol. wt. 84,000 and 27,700 by aqueous-phase gel-permeation chromatography. The fractions contain d-galactose (65 mol%), 4-O-methyl-d-glucuronic acid (24 mol%), d-glucuronic acid (3 mol%), l-arabinose (4 mol%), and l-rhamnose (3 mol%). Smith degradation of the carboxyl-reduced polysaccharides gives products of halved molecular weight, and these consist of a β-(1→3)-linked galactopyranan main chain and side chains made up of galactopyranose residues. Peripheral groups, such as α-d-Galp-, α-d-Galp-(1→6)-β-d-Galp-, 4-O-methyl-β-d-GlcpA-, and 4-O-methyl-β-d-GlcpA-(1→6)-β-d-Galp-, are attached to this interior core through β-(1→3)- or β-(1→6)-linkages.     

Substrate specificities and kinetic properties of two (1→3), (1→4)-β-d-glucan endo-hydrolases from germinating barley (Hordeum vulgare)

Two β-d-glucan endo-hydrolases purified from germinating barley (Hordeum vulgare) hydrolyse (1→4)-β linkages in (1→3),(1→4)-β-d-glucans where the d-glucosyl residue is substituted at O-3, but will not hydrolyse (1→3)-β-d-glucans or (1→4)-β-d-glucans. Methylation analysis of hydrolytic products released from barley (1→3),(1→4)-β-d-glucan indicates that 3-O-β-cellobiosyl-d-glucose and 3-O-β-cellotriosyl-d-glucose are the major oligomers formed. The enzymes exhibit characteristic endo-hydrolase action-patterns on this substrate. Both enzyme can therefore be classified as (1→3),(1→4)-β-d-glucan 4-glucanohydrolases (EC 3.2.1.73). The reduced, pneumococcal polysaccharide RS III, which consists of alternating (1→3)- and (1→4)-linked β-d-glucosyl residues, is hydrolysed by the enzymes to release laminaribiose as a major oligomeric product. Although the kinetic parameters of the two enzymes are similar, one hydrolyses barley (1→3),(1→4)-β-d-glucan at a significantly higher rate than the other and is more stable at elevated temperatures.     

Bis(2-acetamido-3,4,6-tri-O-acetyl-2-deoxy-β-D-glucopyranosyl)amine obtained by a fusion reaction

A unique, alkali-soluble polysaccharide has been isolated from the cell walls of the basidiomycete Coprinus macrorhizus microsporus. The polysaccharide, which is primarily a glucan, contains a large proportion of α-(1→4)-linked d-glucose residues and a smaller amount of β-(1→3) and (1→6) linkages, as suggested by methylation, partial acid hydrolysis, periodate oxidation, and enzymic studies. Hydrolysis of the methylated polysaccharide gave equimolar amounts of 2,4-di- and 2,3-di-O-methyl-d-glucose; no 2,6-di-O-methyl-d-glucose was identified, indicating the absence of branch points joined through O-1, O-3, and O-4. The isolation and identification of 2-O-α- glucopyranosylerythritol from the periodate-oxidized polysaccharide suggests that segments of the a-(1→4)-linked d-glucose residues are joined by single (1→3)-linkages. An extracellular enzyme-preparation from Sporotrichum dimorphosporum (QM 806) containing both β-(1→3)- and α-(1→4)-d-glucanohydrolase activity released 76% of the reducing groups from the polysaccharide. The polysaccharide also contains minor proportions of xylose, mannose, 2-amino-2-deoxyglucose, and amino acids.     

The preparation of a crystalline guanosine trialcohol and its sodium salt

A polysaccharide has been extracted from Cassia corymbosa seeds with cold, acidulated water, and purified to give a water-soluble product containing d-galactose and d-mannose in 4:7 molar ratio. Acid-catalyzed fragmentation, periodate oxidation, methylation, and enzymic hydrolysis showed that the seed gum has a branched structure consisting of a linear chain of β-d-(1→4)-linked mannopyranosyl units, some of which are substituted at O-6 by two α-d-(1→6) galactopyranosyl units mutually linked glycosidically. Methylation analysis of the galactomannan afforded 2,3,4-tri- and 2,3,4,6-tetra-O-methylgalactose, along with 2,3-di- and 2,3,6-tri-O-methylmannose, in the molar ratios of 2:2:2:5. Both the methylation and the periodate-oxidation studies showed ～36.4% of end groups. The significance of these results, together with the findings of partial hydrolysis with acid, are discussed, in relation to ascertaining the structure of the repeating unit of the polysaccharide.     

The crystal structure of methyl 3,4-O-isopropylidene-2,6-di-O-(2,3,4,6-tetra-O-acetyl-β-d-galactopyranosyl)-α-d-galactopyranoside at 97 K

The crystal structure of methyl 3,4-O-isopropylidene-2,6-di-O-(2,3,4,6-tetra-O-acetyl-β-d-galactopyranosyl)-α-d-galactopyranoside (1), C₃₈H₅₄O₂₄ · (C₄H₈O₂)_0.32 was determined by X-ray diffraction;1 crystallises in space group P2₁ with a = 12.480(3), b = 8.821(3), c = 21.182(4)Å, β = 98.46(3)°, and Z = 2. The structure was solved by Patterson-search and Fourier-recycling procedures and refined to R_w(R) = 0.048(0.063), using 4348 [3112 with I> 2σ(I)] independent reflections. The β-d-galactosyl rings are slightly distorted and, due to the isopropylidene group, the α-d-galactoside ring is severely distorted. The conformation near the β-(1→6) and β-(1→2) linkages between the pyranoid rings is not significantly affected by the acetyl groups, but the anomeric C-O-C bridge angles have unusual values. The C-6O-6 bond in the β-d-galactosyl group (1→2)-linked to the α-d-galactoside residue has an unusual gauche—trans conformation with respect to C-4 and O-5. The CH₃-(C = O)-O-C moieties are planar within 0.01Å, and 32.6% of all unit cells contain a molecule of ethyl acetate.     

Structure and biodegradation of the polysaccharide components of egg masses isolated from snails of an Ampullarius species

A polysaccharide preparation, isolated from egg masses deposited by snails of an Ampullarius species, was purified via precipitation with Cetavlon in the presence of sodium borate, and found to contain d-galactose and a smaller proportion of d-glucose, and to have two components with sedimentation coefficients of 10S and 40S. A polysaccharide, isolated from freshly laid egg masses, was highly branched and was shown to contain nonreducing α-d-glucopyranosyl and β-d-galactopyranosyl end-groups, and 3,6-di-O-substituted β-d-galactopyranosyl residues. One or more of the polysaccharide components was a d-glucopyrano-d-galactopyranan with non-reducing α-d-glucopyranosyl end-groups (1→4)-linked to β-d-galactopyranosyl residues. The polysaccharide preparations, obtained from freshly laid egg masses and from those that were left for 10 and 15 days after being laid, were structurally different from each other. With the passage of time, progressive diminution of the 10S component and the proportion of d-glucose in the polysaccharide took place, suggesting that each constituent was consumed preferentially by the snail embryos as an energy source.     

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.     

Structure of a new galactomannan from the ascocarps of Cordyceps cicadae shing

A water-soluble galactomannan (C-3), [α]_D²⁰ +30°, isolated from the rod-like ascocarps of Cordyceps cicadae, was determined to be homogeneous, and the molecular weight was estimated by gel filtration to be 27,000. The polysaccharide is composed of d-mannose and d-galactose in the molar ratio of 4:3. The results of methylation analysis, Smith degradation, stepwise hydrolysis with acid, and ¹³C-n.m.r. spectroscopy indicated that the polysaccharide is of highly branched structure, and composed of α-d-(1→2)-linked and α-d-(1→6)-linked mannopyranosyl residues in the core; some of these residues are substituted at O-6 and O-2 with terminal β-d-galactofuranosyl and α-d-mannopyranosyl groups, and with short chains of β-d-(1→2)-linked d-galactofuranosyl units.     

Galactans from Cryptonemia species. Part II: Studies on the system of galactans of Cryptonemia seminervis (Halymeniales) and on the structure of major fractions

Cryptonemia seminervis biosynthesizes a family of d,l-hybrid galactans based on the classical 3-linked β-d-galactopyranosyl→4-linked α-d- and α-l-galactopyranosyl alternating sequence (A-units→B-units) with major amounts of α-d- and α-l-galactose and 3,6-anhydro-d- and l-galactose and lesser percentages of 3,6-anhydro-2-O-methyl-l-galactose, 2-O-methyl-, 4-O-methyl- and 6-O-methylgalactoses. The dispersion of structures in this family is based on five structural factors, namely: (a) the amount and position of substituent groups as sulfate (major), pyruvic acid ketals, methoxyl and glycosyl side-chain (4-O-methyl galactopyranosyl and/or xylosyl); (b) the ratio galactose/3,6-anhydrogalactose in the B-units; (c) the ratio d,l-galactoses and d,l-3,6-anhydrogalactoses also in the B-units, (d) the formation of diads and (e) the sequence of the diads in the linear backbone. Considering these variables it is not unexpected to find in the fractions studied at least 18 structural units producing highly complex structures. Structural studies carried out in two major fractions (S2S-3 and S2S-4) showed that these galactans were formed mainly by β-d-galactopyranosyl 2-sulfate (20 and 11.9 mol %), β-d-galactopyranosyl 2-sulfate 4,6-O-(1′-carboxyethylidene) (8.9 and 6.0 mol %) and β-d-galactopyranosyl 2,6-sulfate (5.4 and 18.6 mol %), together with 3,6-anhydro-α-l-galactopyranosyl (11.4 and 7.3 mol %) and 3,6-anhydro-α-l-galactopyranosyl 2-sulfate (4.9 and 15.4 mol %) and minor quantities of 12-15 other structural units.Preparative alkaline treatment carried out on fraction (S2S-3) produced a quantitative formation of 3,6-anhydro α-l-galactopyranosyl units from precursor units (α-l-galactose 6-sulfate and α-l-galactose 2,6-sulfate). Kinetic studies on this 3,6-anhydro cyclization show a rate constant of 5.2 × 10⁴ s⁻¹ indicating diads of the type G→L6S/2,6S. Data from chemical, spectroscopic and kinetic studies suggest that, in S2S-3, the agaran block in the d,l-hybrid galactan is composed of the following diads: G(6R)→L6S/2,6S and G2S(P)(2,6S)→LA(2S)(2R)(2M) and the carrageenan block of G2S(P)→D(2S)(2,3S)(3S)(3,6S) in a molar ratio of agaran to carrageenan structures of ∼2:1.     

The chemical synthesis of polysaccharides : Part III. Synthesis of 6-O-α-D-mannopyranosyl-D-mannose and 4-O-α-D-mannopyranosyl-D-mannose. Isolation of A (1→6)-linked D-mannan

When 1,2,3,4-tetra-O-acetyl-α-D-mannopyranose was fused with a catalytic amount of toluene-p-sulphonic acid, 6-O-α-D-mannopyranosyl-D-mannose and 4-O-α-D-mannopyranosyl-D-mannose were isolated after deacetylation of the reaction mixture. No β-D-linked disaccharide was detected in the reaction mixture. When the corresponding β-D-tetra-acetate was fused with zinc chloride as catalyst, higher oligomers were formed, and a D-mannan was isolated and shown to be mainly an α-(1→6)-linked polymer having d.p. of 10. With 5% of zinc chloride, the α-D-tetra-acetate showed oligosaccharide formation, and yielded a smaller proportion of a (1→6)-linked D-mannan.     

Some structural features of the d-glucan from the seed of Mirabilis jalapa

The cotyledon of the seed of Mirabilis jalapa was found to contain a d-glucan. Methylation, periodate oxidation, and graded and enzymic hydrolysis studies were conducted to elucidate its structure. For every 38 d-glucosyl residues therein, 34 are (1→4)- and 3 are (1→3)-linked; the d-glucosyl unit at the branch point is linked through O-1, O-2, and O-4. In some places in the chain, there are at least three (1→3)-linked d-glucosyl residues in a sequence. Both α- and β-d-glucosidic linkages are present in the polysaccharide, the former preponderating. The d-glucan gave with iodine a faint blue color that had λ_max 420 nm.     

Synthesis of two isomeric methyl β-D-xylotriosides containing a (1→2)-β-linkage

Partial hydrolysis with acid, methylation analysis (including uronic acid degradation), Smith degradation, and p.m.r. spectroscopy have been used to determine the primary structure of the capsular polysaccharide of Klebsiella serotype k64. The hexasaccharide repeating-unit, which also contains one O-acetyl substituent, comprises a 4)-α-d-GlcpA-(1 → 3)-α-d-Manp-(1 → 3)-β-d-Glcp-(1 → 4)-α-d-Manp-(1 → chain with a 4,6-O-(l-carboxyethylidene)-β-d-glucopyranosyl and an l-rhamnosyl group attached to the 4-linked d-mannosyl residue at O-2 and O-3, respectively.     

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.     

Synthese der T-antigenen trisaccharide O-β-d-galactopyranosyl-(1→3)-O-(2-acetamido-2-desoxy-α-d-galactopyranosyl)-(1→6)-d-galactopyranose und O-β-d-galactopyranosyl-(1→3)-O-(2-acetamido-2-desoxy-α-d-galactopyranosyl)-(1→6)-d-glucopyranose und deren anknüpfung an proteine

The synthesis of the trisaccharides O-β-d-galactopyranosyl-(1→3)-O-(2-acetamido-2-deoxy-α-d-galactopyranosyl)-(1→6)-d-galactopyranose (15) and O-β-d-galactopyranosyl-(1→3)-O-(2-acetamido-2-deoxy-α-d-galactopyranosyl)-(1→6)-d-glucopyranose (27) is described and the synthesis of α-d-glycosides by reaction of 3,4,6-tri-O-acetyl-2-azido-2-deoxy-β-d-galactopyranosyl chloride with highly reactive hydroxyl groups is discussed. The trisaccharide 27 was coupled with serum albumin by formation of an imine intermediate and reduced to an amine, to yield a synthetic T-antigen. A similar coupling of 15 was unsuccessful.     

Distribution of D-galactosyl groups in guaran and locust-bean gum

Distribution of α-d-galactopyranosyl side-chain groups in two galactomannans, guaran and locust-bean gum, was determined by measurement of the O-acetyl-O-methyl-d-mannitol derivatives obtained from the corresponding primary C-p-tolylsulfonyl polysaccharide derivatives. The O-acetyl-O-methyl-d-mannitol derivatives were produced by β-elimination and methylation, with sodium (methylsulfinyl)methide and methyl iodide, of the primary C-p-toluenesulfinylated galactomannans, followed by sequential acid hydrolysis, reduction, and acetylation of the partially degraded p-tolyl sulfones. The results indicated that side-chain units of guaran are alternately disposed along the d-mannan backbone, whereas those of locust-bean gum are disposed in uniform blocks along the backbone.     

New steroidal saponins of Agave americana

Two new saponins, agavasaponin E and agavasaponin H have been isolated from the methanolic extract of Agave americana leaves and their structures elucidated. Agavasaponin E is 3-O-[β-d-xylopyranosyl-(1→2glc1)-α-l-rhamnopyranosyl-(1→4)-α-l-rhamnopyranosyl-(1→3glc 1)-β-d-glucopyranosyl-(1→4)-β-d-glucopyranosyl-(1→4)-α-d-galactopyranosyl]-(25R)-5α-spirostan-12-on-3β-ol, whereas agavasaponin H is 3-O-[β-d-xylopyranosyl-(1→2 glc 1)-α-l-rhamnopyranosyl-(1→4)-α-l-rhamnopyranosyl-(1→3 glc 1)-β-d-glucopyranosyl-(1→4)-β-d-glucopyranosyl-(1→4)-β-d-galactopyranosyl]-26-O-[β-d-glucopyranosyl]-(25R)-5α-furostan-12-on-3β,22α,26-triol.     

Polysaccharides of basidiomycetes. Alkali-soluble polysaccharides from the mycelium of white rot fungus <Emphasis Type="Italic">Ganoderma lucidum</Emphasis> (Curt.: Fr.) P. Karst

Two polysaccharides were isolated from submergedly cultured mycelium of the basidiomycete Ganoderma lucidum by extraction with alkali followed by fractionation with Fehling reagent. The polysaccharides were shown to be a linear (1→3)-α-D-glucan and a highly branched xylomannan containing a backbone built up of (1→3)-linked α-D-mannopyranose residues, the majority of which are substituted at O-4 by single β-D-xylopyranose residues or by disaccharide fragments β-D-Manp-(1→3)-β-D-Xylp-(1→. Polysaccharide structures were elucidated by NMR spectroscopy in combination with methylation analysis and periodate oxidation. An interesting feature of the xylomannan is the simultaneous presence of α-D-mannopyranose and β-D-mannopyranose residues, the first forming the backbone, and the second being the non-reducing terminal units of disaccharide side chains.