Structure of a neutral exopolysaccharide produced by Lactobacillus delbrueckii ssp. bulgaricus LBB.B26

The neutral exopolysaccharide produced by Lactobacillus delbrueckii ssp. bulgaricus LBB.B26 in skimmed milk was found to be composed of d-glucose and d-galactose in a molar ratio of 2:3. Linkage analysis and 1D/2D NMR ((1)H and (13)C) studies performed on the native polysaccharide, and on an oligosaccharide obtained from a partial acid hydrolysate of the native polysaccharide, showed the polysaccharide to consist of branched pentasaccharide repeating units with the following structure. [structure: see text]     

Characterization of the exopolysaccharide produced by Streptococcus thermophilus 8S containing an open chain nononic acid.

The exopolysaccharide produced by Streptococcus thermophilus 8S in reconstituted skimmed milk is a heteropolysaccharide containing d-galactose, d-glucose, d-ribose, and N-acetyl-d-galactosamine in a molar ratio of 2 : 1 : 1 : 1. Furthermore, the polysaccharide contains one equivalent of a novel open chain nononic acid constituent, 3,9-dideoxy-d-threo-d-altro-nononic acid, ether-linked via C-2 to C-6 of an additional d-glucose per repeating unit. Methylation analysis and 1D/2D NMR studies (1H and 13C) performed on the native polysaccharide, and mass spectrometric and NMR analyses of the oligosaccharide obtained from the polysaccharide by de-N-acetylation followed by deamination and reduction demonstrated the 'hepta'saccharide repeating unit to be: -->4)-alpha-D-Galp-(1-->2)-beta-D-Ribf-(1-->4)-beta-D-Galp-(1-->4)-beta-D-Glcp-(1--7')-Sub-(1-->4)-beta-D-GalpNAc-(1--> in which Sug is 6-O-(3',9'-dideoxy-d-threo-d-altro-nononic acid-2'-yl)-alpha-d-glucopyranose.     

Structure of the exopolysaccharide produced by Streptococcus thermophilus S3

The exopolysaccharide of Streptococcus thermophilus S3, produced in skimmed milk, is composed of D-galactose and L-rhamnose in a molar ratio of 2:1. The polysaccharide contains 0.4 equiv of O-acetyl groups per repeating unit. Linkage analysis and 1D/2D NMR (1H and 13C) studies on native and O-deacetylated EPS together with nanoES-CID tandem mass spectrometry studies on oligosaccharides generated by a periodate oxidation protocol, show the polysaccharide to have the following structure: [structure: see text].     

Structural determination of a neutral exopolysaccharide produced by Lactobacillus delbrueckii ssp. bulgaricus LBB.B332

The neutral exopolysaccharide produced by Lactobacillus delbrueckii ssp. bulgaricus LBB.B332 in skimmed milk was found to be composed of d-glucose, d-galactose, and l-rhamnose in a molar ratio of 1:2:2. Linkage analysis and 1D/2D NMR (1H and 13C) studies carried out on the native polysaccharide as well as on an oligosaccharide generated by a periodate oxidation protocol, showed the polysaccharide to consist of linear pentasaccharide repeating units with the following structure: -->3-alpha-D-Glcp-(1-->3)-alpha-D-Galp-(1-->3)-alpha-L-Rhap-(1-->2)-alpha-L-Rhap-(1-->2)-alpha-D-Galp-(1-->.     

The structure of the specific capsular polysaccharide of Rhodococcus equi serotype 4

The specific capsular polysaccharide produced by Rhodococcus equi serotype 4 was found to be a high-molecular-weight acidic polymer composed of D-glucose, D-mannose, pyruvic acid and a previously unidentified 5-amino-3,5-dideoxynonulosonic (rhodaminic) acid in the proportions 2:1:1:1. Structural analysis, employing a combination of microanalytical methods, nuclear magnetic resonance spectroscopy, and mass spectrometric techniques, established that the polysaccharide consisted of linear repeating tetrasaccharide units having the sequence of residues shown below. In the native polysaccharide, the rhodaminic acid residues were present as their acetamido derivatives (RhoANAc) and carried 1-carboxyethylidene groups that bridged the O-7 and O-9 positions. Treatment of the capsular polysaccharide with dilute acetic acid and/or anhydrous hydrogen fluoride under hydrolytic/solvolytic conditions, resulted in the formation of four different oligosaccharide species. The 1H and 13C NMR resonances of these oligosaccharide fragments and of the native serotype 4 capsular polysaccharides were fully assigned by homo- and heteronuclear chemical shift correlation methods.     

Structure of the capsular polysaccharide of Klebsiella serotype K40

The capsular polysaccharide from Klebsiella Serotype K40 contains D-galactose, D-mannose, L-rhamnose, and D-glucuronic acid in the ratios of 4:1:1:1. Methylation analysis of the native and carboxyl-reduced polysaccharide provided information about the glycosidic linkages in the repeating unit. Degradation of the permethylated polymer with base established the identity of the sugar unit preceding the glycosyluronic acid residue. The modes of linkages of different sugar residues were further confirmed by Smith degradation and partial hydrolysis of the K40 polysaccharide. The anomeric configurations of the different sugar residues were determined by oxidation of the peracetylated native and carboxyl-reduced polysaccharide with chromium trioxide. Based on all of these results, the heptasaccharide structure 1 was assigned to the repeating unit of the K40 polysaccharide. (Formula: see text)     

Structural characterization and immunomodulating activity of a complex glucan from spores of Ganoderma lucidum.

A polysaccharide with a molecular weight of 1.26 x 10(5), obtained from the sporoderm-broken spores of Ganoderma lucidum, was purified by anion-exchange and gel-permeation chromatography. This polysaccharide had a strong effect on suppressing the antibody production and the Con A or LPS induced lymphocyte proliferation in mice. Chemically, the structure of the polysaccharide was identified by methylation analysis, 1 D, 2 D NMR and ESI-MS spectroscopic studies of the native one and of the oligosaccharide fragments generated by partial acid hydrolysis, Smith degradation, and acetolysis. It was concluded that the intact polysaccharide was a complex beta-D-glucan consisting of a (1-->6)-linked backbone chain, in which every other glucosyl residue was substituted at C-3 or C-4 with mono-, di- and trisaccharide branches.     

Structural Characterization and Immunomodulating Activity of a Complex Glucan from Spores of Ganoderma lucidum

A polysaccharide with a molecular weight of 1.26×10⁵, obtained from the sporoderm-broken spores of Ganoderma lucidum, was purified by anion-exchange and gel-permeation chromatography. This polysaccharide had a strong effect on suppressing the antibody production and the Con A or LPS induced lymphocyte proliferation in mice. Chemically, the structure of the polysaccharide was identified by methylation analysis, 1D, 2D NMR and ESI-MS spectroscopic studies of the native one and of the oligosaccharide fragments generated by partial acid hydrolysis, Smith degradation, and acetolysis. It was concluded that the intact polysaccharide was a complex β-D-glucan consisting of a (1→6)-linked backbone chain, in which every other glucosyl residue was substituted at C-3 or C-4 with mono-, di- and trisaccharide branches.     

Structural characterisation and enzymic modification of the exopolysaccharide produced by Lactococcus lactis subsp. cremoris B39

Lactococcus lactis subsp. cremoris B39 grown on whey permeate produced an exopolysaccharide containing L-Rha, D-Gal and D-Glc in a molar ratio of 2:3:2. The polysaccharide was modified using an enzyme preparation from Aspergillus aculeatus, resulting in the release of Gal and a polymer with approximately the same hydrodynamic volume as the native polysaccharide. Linkage analysis and 1H NMR studies of both the native and modified exopolysaccharides elucidated that terminally linked Gal was released during modification and that the chemical structure of the branches within the repeating units is: beta-D-Galp-(1-->4)-beta-D-Glcp-(1-->. 2D NMR experiments (both 1H-1H and 1H-13C) revealed that exopolysaccharide B39 consists of a branched heptasaccharide repeating unit with the following structure: [structure: see text].     

Structural determination and immunochemical characterization of the type V group B Streptococcus capsular polysaccharide

The type V capsular polysaccharide of group B Streptococcus has been isolated and purified, and its repeating unit structure determined. The native type V polysaccharide contains D-glucose, D-galactose, 2-acetamido-2-deoxy-D-glucose, and sialic acid in a molar ratio of 3:2:1:1. Methylation analysis and 1H NMR and 13C NMR analysis of the native type V polysaccharide and of its specifically degraded products permitted the determination of the repeating unit structure of the type V polysaccharide: [formula: see text] The type V polysaccharide has certain structural features in common with other group B streptococcal capsular polysaccharides but is antigenically distinct: no immunologic cross-reactivity was observed between type V and types Ia, Ib, II, III, or IV polysaccharides. Studies of antibody binding to the partially degraded forms of the type V polysaccharide indicated that the native epitope is complex, involving most if not all of the sugar residues of the repeating unit.     

Isolation and structural elucidation of a water-soluble polysaccharide (PS-I) of a wild edible mushroom, Termitomyces striatus

A heteropolysaccharide (PS-I), isolated from the hot aqueous extract of an edible mushroom, Termitomyces striatus, is composed of d-glucose, d-galactose, d-mannose and l-fucose in a molar ratio 2:1:1:1. Structural investigation of the native as well as the Smith-degraded polysaccharide was carried out using methylation analysis, periodate oxidation studies and 1D and 2D NMR spectroscopy, and the repeating unit of the polysaccharide is established as follows: [carbohydrate structure: see text]     

Bacterial antigenic polysaccharides. 12. Structure and 13C NMR spectrum of the polysaccharide chain of Shigella boydii type 8 lipopolysaccharide

A specific acidic polysaccharide was isolated from Sh. boydii type 8 antigenic lipopolysaccharide after mild hydrolysis followed by chromatography on Sephadex G-50. The polysaccharide consists of D-glucuronic acid, D-galacturonic acid, 2-acetamido-2-deoxy-D-glucose, 2-acetamido-2-deoxy-D-galactose and 2-amino-1,3-propanediol residues in 1:1:1:1:1 ratio. From the results of methylation analysis, partial acid hydrolysis and Smith degradation, the structure of the repeating unit of the specific polysaccharide was deduced as: (Formula: see text). The 13C NMR spectra of native, O-deacetylated and carboxyl-reduced polysaccharides, as well as the spectrum of oligosaccharide produced by Smith degradation were interpreted. The 13C NMR data fully confirmed the structure of the polysaccharide repeating unit.     

Antigenic polysaccharides of bacteria of the genus Shigella. The structure of the polysaccharide chain of Shigella boydii type 14

A specific acidic polysaccharide has been isolated from the Shigella boydii type 14 antigenic lipopolysaccharide after mild hydrolysis followed by chromatography on Sephadex G-50. The polysaccharide consists of the D-glucuronic acid, 2-acetamido-2-deoxy-D-glucose and D-galactose residues in the ratio 1:1:3. From the results of methylation analysis and partial acid hydrolysis, the structure of the repeating unit of the specific polysaccharide was deduced as follows: (-6DGalp alpha 1-4DGlcAp beta 1-6DGalp beta 1-4DGalp beta 1-4DGlcNAcp beta 1-)n. The 13C NMR spectra of native and carboxyl-reduced polysaccharides, as well as of oligosaccharides produced by partial acid hydrolysis fully confirmed the proposed structure. The approach was suggested to determine the type of substitution of uronic acid moieties in polysaccharide chain by use of chromato-mass-spectrometry of acetylated methyl esters of partially methylated aldonic acids. Serological characteristics of Sh. boydii LPS type 14 and its modified derivatives are discussed.     

Somatic antigens of Shigella. Structural investigation on the O-specific polysaccharide chain of Shigella dysenteriae type-2 lipopolysaccharide.

The O-specific polysaccharide obtained from Shigella dysenteriae type-2 lipopolysaccharide by mild acid hydrolysis consisted of N-acetylgalactosamine, N-acetylglucosamine, D-galactose, D-glucose, and O-acetyl group in the ratio of 2:1:1:1:1. A number of oligosaccharides were obtained by deamination of the N-deacetylated polysaccharide and by Smith degradation of the both native and O-deacetylated polysaccharides. The identification of oligosaccharides along with methylation analysis and chromic anhydride oxidation showed that the polysaccharide was built up of the repeating pentasaccharide units whose proposed structure is given below: (see article) Serological properties of Sh. dysenteriae O-specific polysaccharides are discussed.     

Structural and immunological studies of a major polysaccharide from spores of Ganoderma lucidum (Fr.) Karst

A polysaccharide isolated from spores of the fungus, Ganoderma lucidum, was found to be a complex glucan. On the basis of compositional and methylation analyses, periodate oxidation, Smith degradation, 1D and 2D NMR, and ESIMS experiments of the native polysaccharide and its degraded products, the polysaccharide was shown to have a backbone of beta-(1-->3)-linked D-glucopyranosyl residues, with branches of mono-, di- and oligosaccharide side chains substituting at the C-6 of the glucosyl residues in the main chain. Conformational analysis in aqueous solution and immunological activities of the native and degraded glucans were also investigated. The results suggested that the degree of substitution on the main chain and the length of side chains may be very important factors in determining the conformation and the biological activities of beta-(1-->3)-linked glucans.     

Chemical characterization of capsular polysaccharide from Cryptococcus neoformans serotype A-D

During a study of serotyping of Cryptococcus neoformans, we found that the type strain of C. neoformans (CBS 132) was serotype A-D. This strain agglutinated with both factor 7 serum (specific for serotype A) and factor 8 serum (specific for serotype D) in our serotyping system. Therefore, we investigated the chemical structure of the antigenic capsular polysaccharide of this strain. The soluble capsular polysaccharide was obtained from the culture supernatant fluid by precipitation with ethanol. Column chromatography of the polysaccharide on DEAE-cellulose yielded three fractions (F-1 to F-3). The major antigenic activity was found in the F-3 fraction. The results obtained by methylation analysis, controlled Smith degradation-methylation analysis, partial acid hydrolysis, and other structural studies of F-3 polysaccharide indicated that the polysaccharide contains mannose, xylose, and glucuronic acid at a ratio of 7:2:2, and has a backbone of alpha (1-3)-linked D-mannopyranoside residues with a single branch of beta (1-2)-xylose and glucuronic acid. The ratio of mannose residues with or without a branch in the F-3 polysaccharide was 4:3 and its molecular weight calculated from the average of the degree of polymerization was 46,500 daltons. These results indicate that the chemical structure of the capsular polysaccharide of serotype A-D is very similar to those from serotypes A and D, suggesting that small differences in the molar ratio and pattern of linkage of monosaccharides in the branch of the polysaccharides of the three serotypes may be responsible for their different specificities.     

Structure of the capsular polysaccharide of Escherichia coli O9:K32(A):H19

The capsular polysaccharide of the bacterium Escherichia coli O9:K32(A):H19 was analyzed using chemical methods (hydrolysis, sequential Smith degradation, methylation analysis) together with 1H- and 13C-n.m.r. spectroscopy. 13C-N.m.r. spectroscopy and chemical analyses indicated that the K32 polysaccharide is composed of equimolar proportions of glucose, galactose, rhamnose, and glucuronic acid, and carries O-acetyl groups. 1H-N.m.r. analysis of native K32 polysaccharide revealed five resonances in the anomeric region (delta 5.52, 5.16, 5.12, 5.02, and 4.73) and the presence of an acetyl group (delta 2.18). O-Deacetylation of the polysaccharide resulted in the loss of the resonance at delta 2.18 and one of the resonances (delta 5.52) in the anomeric region. The "extra" anomeric resonance in the 1H-n.m.r. spectrum of the native K32 polymer was assigned to H-2 of rhamnose, which experiences a large downfield shift when the 2-position is O-acetylated. This was confirmed by a 2D-COSY n.m.r. experiment and studies of model compounds. The K32 capsular polysaccharide is of the "2 + 2" type, comprised of the following repeating unit: (sequence; see text) This structure is identical to that of Klebsiella K55 capsular polysaccharide.     

Structure of a highly pyruvylated galactan sulfate from the Pacific green alga Codium yezoense (Bryopsidales, Chlorophyta)

A polysaccharide fraction consisting of d-galactose, sulfate, and pyruvate in a molar proportion of 4:2:1 was isolated from the green seaweed Codium yezoense by water extraction followed by ion-exchange chromatography. To elucidate its structure, modified polysaccharides were prepared by desulfation, depyruvylation, and by total removal of non-carbohydrate substituents. Structures of the native polysaccharide and of the products of its chemical modifications were investigated by methylation analysis as well as by 1D and 2D (1)H and (13)C NMR spectroscopy. The polysaccharide devoid of sulfate and pyruvate was subjected to two subsequent Smith degradations to afford a rather low-molecular and essentially linear (1-->3)-beta-d-galactan. A highly ramified structure was suggested for the native polysaccharide, which contains linear backbone segments of 3-linked beta-d-galactopyranose residues connected by (1-->6) linkages, about 40% of 3-linked residues being additionally substituted at C-6, probably by short oligosaccharide residues also containing (1-->3) and (1-->6) linkages. Sulfate groups were found mainly at C-4 and in minor amounts at C-6. Pyruvate was found to form mainly five-membered cyclic ketals with O-3 and O-4 of the non-reducing terminal galactose residues. The minor part of pyruvate forms six-membered cyclic ketals with O-4 and O-6. The absolute configurations of ketals (R for six-membered ketals and S for five-membered ones) were established using NMR spectral data.     

Structure elucidation of the O-antigenic polysaccharide from the enteroaggregative Escherichia coli strain 62D1.

The O-antigen polysaccharide of the lipopolysaccharide from the enteroaggregative Escherichia coli strain 62D1 has been determined. Sugar and methylation analysis together with 1H and 13C NMR spectroscopy revealed the components of the repeating unit. Two-dimensional NOESY and heteronuclear multiple-bond correlation experiments were used to deduce the sequence. 1H and 13C NMR spectra indicate heterogeneity in the polysaccharide. Methylation analysis and 1H NMR spectra of native and Smith-degraded material show that the majority (65%) of the repeating units has the following structure: Minor resonances in the NMR spectra are consistent with the presence of repeating units which lack the alpha-d-Galp terminal residue (35%).     

Chemical structure of an acidic polysaccharide produced by serratia piscatorum

The acidic polysaccharide of Serratia piscatorum consists of L-rhamnopyranosyl, D-galactopyranosyl, and D-galactopyranosyluronic acid residues in the molar ratio of 2:1:1. Some of the D-galactopyranosyluronic acid residues are acetylated at O-2 or O-3, or both. Smith degradation and methylation analysis indicated that the L-rhamnopyranosyl, D-galactopyranosyl, and D-galactopyranosyluronic acid residues are substituted with glycosidic linkages at O-3, O-3, and O-4, respectively. Partial acid hydrolysis of the native polysaccharide gave four acidic oligosaccharides, each of which was isolated and characterized, suggesting the following tetrasaccharide repeating unit: →3)-L-Rhap-(1→4)-D-GalAp-(1→3)-L-Rhap-(1→3)-D-Galp-(1→.