Capsular and extracellular polysaccharides from Rhizobium microsymbionts of Acacia decurrens

The capsular polysaccharide produced by a Rhizobium isolated from a root nodule of Acacia decurrens is composed of 3-O-methyl- -rhamnose: -rhamnose: - mannose: -glucose: -galacturonic acid in the molar ratios of 1:2:2:4:1. The extracellular polysaccharide is similarly constituted. Structural analyses indicate a decasaccharide repeating-unit in which the -rhamnosyl groups occur as single-unit side-chains. The 3-O-methyl- -rhamnosyl and one of the α- -rhamnosyl groups are (1→6)-linked to two of the -glucosyl residues. The other α- -rhamnosyl group is (1→4)-linked to the -galacturonic acid residue. The main-chain residues are all (1→3)-linked, and are partially identified as -(1→3)-α- -GalpA-(1→3)-α- -Manp- (1→3)-α- -Glcp-(1→3)-.     

Structural studies on hairy region of pectic polysaccharide from campion Silene vulgaris (Oberna behen)

Using enzymic digestion with pectinase, controlled Smith degradation and NMR-spectroscopy, some structural features of the hairy region of pectic polysaccharide termed silenan SV from the aerial part of campion Silene vulgaris (Moench) Garke (Oberna behen (L.) Ikonn) were elucidated.

Silenan was subjected to enzymic digestion with pectinase to furnish the polysaccharide fraction (SVP). The contained residues of -galacturonic acid (43%), arabinose, galactose and rhamnose as main constituents. The backbone of the hairy region of silenan was found to consist of -1,4-galactopyranosyl uronic acid and 2-O-glycosylated rhamnopyranose residues. The side chains contained linear regions of residues of -1,5-linked arabinofuranose and β-1,3-, β-1,4-linked galactopyranose. Silenan SV and its fragment SVP were subjected to Smith degradation to give fractions SVS and SVPS. These contain the residues of terminal and 2-substituted -arabinofuranose as well as residues of terminal, 3-, and 2,3-substituted β-galactopyranose. In addition, NMR-spectral data confirmed that the residues of -rhamnopyranose 2-O-glycosylated with the residues of -1,4-galactopyranosyl uronic acid of the backbone occurred in the core of SVPS and, therefore, in the backbone of silenan SV.

On the basis of data obtained, the hairy regions of silenan were suggested to contain mainly the linear chains of β-1,3-, β-1,4-galactopyranan and -1,5-arabinofuranan. The chains of -1,5-linked arabinofuranose, β-1,3- and β-1,4-linked galactopyranose were shown to be involved in the side chains of the hairy region having branching points at 2,3-substituted β-galactopyranose residues.     

Detailed Structure of an Acidic Polysaccharide in Soy Sauce,Confirmed by Use of Two Kinds of Purified Pectinases

The partially degraded polysaccharide obtained by means of mild acid hydrolysis of APS–I, an acidic polysaccharide in soy sauce, was incubated with an endo-polygalacturonase, and some acidic sugars liberated were fractionated and purified from the enzymatic hydrolyzate. They were identified as d-galacturonic acid (Gal A), its α-l,4-linked dimer and trimer, d-xylose (Xyl) β1→3 Gal A and Xyl β1→3 Gal A α1→4 Gal A, and an acidic polymer composed of the above-mentioned sugars through methylation analysis and other methods.

Methyl-esterized APS–I was subjected to β-elimination with pectin lyase or by heat treatment. The high molecular fraction which was highly resistant to β-elimination, was concluded to be β-1,4-linked d-galactan of which reducing-end group was attached to d-galacturonic acid. On the basis of these findings and previous knowledges, the construction of APS–I was also discussed.     

Acid and Enzymatic Hydrolysis of an Acidic Polysaccharide from Soy Sauce

Mild acid hydrolysis of an acidic polysaccharide (APS-I) from soy sauce resulted in a degraded polysaccharide (DPS), the mixture of neutral sugar, D-galacturonic acid, its α-1,4-linked homologous di- and trisaccharides, and acidic oligosaccharides containing residues of D-galacturonic acid and L-rhamnose. Besides the above-mentioned sugars, an aldobiouronic acid containing D-xylose moiety was also yielded in the enzymatic hydrolysates with a crude polysaccharidase preparation. However, only a β-l, 4-galactobiose was isolated from the lower molecular fraction of enzymatic digest of APS-I with a typical hemicellulase preparation. DPS containing 83% of D-galacturonic acid was able to be degraded by endo-polygalacturonase, but APS-I was not because of its highly was discussed on the basis of these results, periodate oxidation study.     

Structural studies and physiological activity of lemnan, a pectin fromLemna minor L

A pectic polysaccharide, lemnan, was isolated from freshly collected duckweedLemna minor L. Its sugar chain was shown to be mainly composed of the residues ofD-galacturonic acid (64%), galactose, arabinose, xylose, andD-apiose, a branched chain sugar. The high content ofD-apiose (25%) indicated that lemnan is an apiogalacturonan type pectin similar to zosteran, a pectic polysaccharide from a sea phanerogam of the Zosteraceae family. The results of partial acidic hydrolysis, pectinase digestion, and NMR studies of lemnan demonstrated that its macromolecule contains regions of the linear α-1,4-D-galacturonan and branched apiogalacturonan. The side chains of apiogalacturonan were found to be formed of single and 1,5-linked residues ofD-apiofuranose attached to 2- and 3-positions of theD-galacturonic acid residues of the apiogalacturonan backbone. Lemnan was shown to exhibit an immunomodulatory effect activating the system of phagocytosis.     

Structural characterization and antioxidant activity of a polysaccharide from the fruiting bodies of cultured Cordyceps militaris

The water-soluble crude polysaccharides were obtained from the fruiting bodies of cultured Cordyceps militaris by hot water extraction followed by ethanol precipitation. The polysaccharides were successively purified by chromatography on DEAE–cellulose-52 and Sephacryl S-100 HR columns, giving main three polysaccharide fractions termed P50-1, P70-1, and P70-2. Structural features of P70-1 were investigated by a combination of chemical and instrumental analysis, such as partial acid hydrolysis, methylation analysis, periodate oxidation – Smith degradation, GC–MS, ¹³C NMR, HPAEC-PAD, and FT-IR. The results indicated that P70-1 has a backbone of (1 → 6)-linked β-d-mannopyranosyl residues, which occasionally branches at O-3. The branches were mainly composed of (1 → 4)-linked -d-glucopyranosyl and (1 → 6)-linked β-d-galactopyranosyl residues, and terminated with β-d-galactopyranosyl residues and -d-glucopyranosyl residues. In the in vitro antioxidant assay, P70-1 was found to possess hydroxyl radical-scavenging activity with an IC₅₀ value of 0.548 mg/ml.     

Characterization of a biologically active arabinogalactan from the leaves of Plantago major L.

PMIa is a Type II arabinogalactan with anti-complementary activity isolated from the leaves of Plantago major L. It has a molecular weight of 77000–80000 Da and consists of arabinose (38%), galactose (49%), rhamnose (6%), galacturonic acid (7%) and 1.5% protein with hydroxyproline, alanine and serine as the main amino acids. Characterization of PMIa by methylation and GC-MS, methanolysis and GC, Smith degradation, weak acid hydrolysis, ¹³C-NMR, ¹H-NMR, two-dimensional heteronuclear NMR and DEPT show that it consists of 1,3-linked galactan chains with 1,6-linked galactan side chains attached to position 6. The side chains are further branched in position 3 with 1,3-linked galactose residues which have 1,6-linked galactose attached to position 6; these 1,3- and 1,6-linked galactose chains altogether probably form a network. Terminal and 1,5-linked arabinose in furanose form are attached to the galactan mainly through position 3 of the 1,6-linked galactose side chains.     

An unusual juxtaposition of polysaccharide components of Collema leptosporum

The lichenised ascomycete, Collema leptosporum Malme, was extracted with aqueous methanol to give traces of mannitol and 3-O-β- -glucopyranosyl- -mannitol (2.7% yield). The residue was consecutively extracted with hot water to give a complex uronic acid-containing polysaccharide, and then with hot aqueous alkali which provided a mixture of polysaccharides. This was fractionated with Cetavlon to give a branched galactomannan, which had the lowest content of galactose yet reported for such a lichen polysaccharide. It has a main chain of (1→6)-linked -Manp units partly substituted at O-2,4 by non-reducing end-units of Manp and Galp, shown by NMR spectroscopy to have - and β-configurations, respectively. The other polysaccharide component was unexpectedly a branched (1→3), (1→6)-linked β-glucan, which is typical of a basidiomycete, whereas those of ascomycetes contain similar linkages but in linear glucans.     

Structure of the serine-containing capsular poly-saccharide K40 antigen from Escherichia coli O8:K40:H9

The structure of the K40 antigenic capsular polysaccharide (K40 antigen) of E. coli O8:K40:H9 was elucidated by determination of the composition, ¹H- and ¹³C-n.m.r. spectroscopy, periodate oxidation and Smith degradation, and methylation analysis. The K40 polysaccharide consists of [(O-β- -glucopyranosyluronic acid)-(1→4)-O-(2-acetamido-2-deoxy-- -glucopyranosyl)-(1→6)-O-(2-acetamido-2-deoxy-- -glucopyranosyl)-(1→4)] repeating units. All of the glucuronic acid residues are substituted amidically with -serine.     

Analysis by the reductive-cleavage method of linkage positions in a polysaccharide containing 4-linked D-glucopyranosyluronic residues

The fate of 4-linked D-glucopyranosyluronic residues under reductive-cleavage conditions was investigated by using the Klebsiella aerogenes type 54 strain A3 capsular polysaccharide. Treatment of the fully methylated polysaccharide with triethylsilane and trimethylsilyl trifluoromethanesulfonate in dichloromethane, followed by in situ acetylation, yielded 1,5-anhydro-2,3,4,6-tetra-O-methyl-D-glucitol, 3,4-di-O-acetyl-1,5-anhydro-2,6-di-O-methyl-D-glucitol, and 3-O-acetyl-1,5-anhydro-2,4-di-O-methyl-L-fucitol, as expected, but the expected product of reductive cleavage of the 4-linked D-glucopyranosyluronic residue, namely, methyl 3-O-acetyl-2,6-anhydro-4,5-di-O-methyl-L-gulonate, was not observed. Instead, methyl 2-O-acetyl-3,6-anhydro-4,5-di-O-methyl-L-gulonate (6) was identified as the sole product of reductive cleavage of the 4-linked D-glucopyranosyluronic residue. That compound 6 arose as a result of rearrangement during reductive cleavage rather than by reductive cleavage of a 5-linked D-glucofuranosyluronic residue, was established by reductive cleavage of the fully methylated polysaccharide following reduction of its ester groups with either lithium aluminum hydride or lithium aluminum deuteride. The products of the latter reductive cleavage were the same as before, except for the absence of 6 and the presence of 4,6-di-O-acetyl-1,5-anhydro-2,3-di-O-methyl-D-glucitol, or its 6,6-dideuterio isomer. Although the reductive-cleavage technique is suitable for the direct analysis of polysaccharides containing 4-linked D-glucopyranosyluronic residues, it does not establish whether the uronic residue is a 4-linked pyranoside or a 5-linked furanoside. The expected product is, however, derived from the 4-linked D-glucopyranosyluronic residue after sequential methylation, reduction of its ester group and reductive cleavage.     

A study of the glucofructofuranan from the New Zealand cabbage tree Cordyline australis

Extraction of the roots of the New Zealand cabbage tree Cordyline australis with water gave a glucofructofuranan in 60% yield (dry-weight basis). Viscosity measurements on aqueous solutions of the polysaccharide, and vapor pressure osmometry of the polysaccharide peracetate, showed the number average molecular weight of the glucofructofuranan to be 3000. Complete hydrolysis with dilute acid gave only -fructose and -glucose, in the ratio of 16:1. The polysaccharide was methylated by using dimethyl sulfoxide—sodium hydroxide—methyl iodide, and the methylated polymer was hydrolyzed to give 1,3,4,6-tetra-O-methylfructose (5.6 mol), 2,3,4,6-tetra-O-methylglucose (1 mol), 1,3,4-tri-O-methylfructose (8.4 mol), 2,3,4-tri-O-methylglucose (0.1 mol), and 3,4-di-O-methylfructose (2.7 mol). These results, supported by ¹³C-n.m.r. analyses, showed that the polymer is a highly branched glucofructofuranan containing mainly (1→2)-linked β- -fructofuranosyl residues, with branching at O-6 of 15% of the -fructosyl residues.     

不同黄腐酸用量对‘红将军’苹果产量、品质和15N-尿素去向的影响

以15年生红将军/八棱海棠为试材,运用¹⁵N同位素示踪技术,设置单施尿素(CK)及尿素配施不同用量黄腐酸处理(黄腐酸用量分别为75、150、300和450 kg·hm^-2,分别以NF₁、NF₂、NF₃和NF₄表示),研究不同黄腐酸用量对苹果植株¹⁵N-尿素吸收、利用、残留、损失及果实产量和品质的影响.结果表明: 至果实成熟期,苹果根系、一年生枝和叶片的Ndff值(植株器官从肥料中吸收分配到的¹⁵N量对该器官全氮量的贡献率)均为NF₃>NF₄>NF₂>NF₁>CK,且各处理间差异显著.植株全氮量和¹⁵N吸收量均以NF₃处理最大,其次为NF₄处理,CK处理最低.与CK处理相比,NF₁、NF₂、NF₃和NF₄处理¹⁵N利用率分别提高了14.2%、33.5%、64.2%和50.0%,而¹⁵N损失率分别降低了9.1%、18.5%、37.1%和28.7%.不同处理土壤¹⁵N残留量不同.配施黄腐酸处理0～60 cm土层¹⁵N残留量显著高于CK处理,其中以NF₃处理最多,而在60～100 cm土层显著低于CK处理.NF₃处理单株产量和纯收益较CK处理增幅最大,分别为15.8%和20.2%,其次为NF₄处理,同时,NF₃处理果实硬度、可溶性固形物含量和糖酸比均达到最高水平.通过对果实产量和氮素利用率与黄腐酸施用量进行拟合分析,得出本试验条件下适宜的黄腐酸用量为326.41～350.61 kg·hm^-2.     

不同黄腐酸用量对‘红将军’苹果产量、品质和15N-尿素去向的影响

以15年生红将军/八棱海棠为试材,运用¹⁵N同位素示踪技术,设置单施尿素(CK)及尿素配施不同用量黄腐酸处理(黄腐酸用量分别为75、150、300和450 kg·hm^-2,分别以NF₁、NF₂、NF₃和NF₄表示),研究不同黄腐酸用量对苹果植株¹⁵N-尿素吸收、利用、残留、损失及果实产量和品质的影响.结果表明: 至果实成熟期,苹果根系、一年生枝和叶片的Ndff值(植株器官从肥料中吸收分配到的¹⁵N量对该器官全氮量的贡献率)均为NF₃>NF₄>NF₂>NF₁>CK,且各处理间差异显著.植株全氮量和¹⁵N吸收量均以NF₃处理最大,其次为NF₄处理,CK处理最低.与CK处理相比,NF₁、NF₂、NF₃和NF₄处理¹⁵N利用率分别提高了14.2%、33.5%、64.2%和50.0%,而¹⁵N损失率分别降低了9.1%、18.5%、37.1%和28.7%.不同处理土壤¹⁵N残留量不同.配施黄腐酸处理0～60 cm土层¹⁵N残留量显著高于CK处理,其中以NF₃处理最多,而在60～100 cm土层显著低于CK处理.NF₃处理单株产量和纯收益较CK处理增幅最大,分别为15.8%和20.2%,其次为NF₄处理,同时,NF₃处理果实硬度、可溶性固形物含量和糖酸比均达到最高水平.通过对果实产量和氮素利用率与黄腐酸施用量进行拟合分析,得出本试验条件下适宜的黄腐酸用量为326.41～350.61 kg·hm^-2.     

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.     

Structural study of bergenan,a pectin from <Emphasis Type="Italic">Bergenia crassifolia</Emphasis>

A pectin polysaccharide named bergenan was isolated from the freshly collected leaves of the leather bergenia Bergenia crassifolia by extraction with an aqueous solution of ammonium oxalate. The main component of its carbohydrate chain was shown to be the residues of D-galacturonic acid (about 80%). In addition, the polysaccharide contains the residues of galactose, arabinose, and rhamnose; their total content is less than 15%. It was shown that the bergenan samples from bergenia leaves collected at different vegetation periods (from July to September) do not substantially differ either in monosaccharide composition or in the viscosity of their aqueous solutions. The results of enzymatic hydrolysis by α-1,4-galacturonase (pectinase), partial acidic hydrolysis, NMR spectroscopy, and methylation with subsequent analysis of the results by GC-MS indicate that the bergenan macromolecule contains the regions of a linear α-1,4-D-galactopyranosyluronan and rhamnogalacturonan-I (RG-I). Galacturonan responds for a greater part of the macromolecule. A considerable amount of its constitutent galacturonic acid residues are present as methyl esters. The side chains in RG-I are attached to the rhamnopyranose residues of the backbone by a 1,4-linkages and are composed of the residues of terminal arabinofuranose and galactopyranose, 1,5-linked α-arabinofuranose, and 1,4- and 1,6-linked β-galactopyranose. The branching points of the side chains of the RG-I molecule are 3,4- and 3,6-di-O-substituted galactose residues.     

O-maleoyl derivative of low-molecular-weight κ-carrageenan: Synthesis and characterization

Low-molecular-weight (LMW) κ-carrageenan was achieved through mild hydrochloric acid hydrolysis of κ-carrageenan. The acylation of low-molecular-weight κ-carrageenan was performed by use of tetrabutylammonium (TBA) salt of the anionic polysaccharide fragments, maleic anhydride, 4-dimethylaminopyridine and tributylamine under homogeneous conditions in N,N-dimethylformamide at 60 °C. Analysis of FT-IR spectrum of O-maleoyl κ-carrageenan showed that a monoester derivative with maleoyl group was formed when LMW κ-carrageenan reacted with maleic anhydride. Investigation of ¹H NMR spectroscopy revealed that the maleoylation took place regioselectively at C-2 of the κ-carrageenan 3-linked unit.     

Sudies of O-specific polysaccharide chains of Pseudomonas solanacearum lipopolysaccharides consisting of structually different repeating units.

The structure of the O-antigenic ploysaccharide chains of lipopolysaccharides of a number of Pseudomonas solanacearum strains were elucidated mainly with the help of methylation analysis and ¹³CNMR spectroscopy, including a computer-assisted ¹³CNMR-based analysis. Six structually distinct but related polysaccharides were identified. They have a backbone which is built up of three -rhammoyranosyl resides and one 2-acetamido-2-deoxy- -glucopyranosyl residue, and is unsubtituted or substituted with a residue of -xylopyranose or -rhamnopyranose as a monosaccharide side chain. The lipopolysaccharides of most of the strains contain polysaccharide chains consisting of at least two structually different types of repeating units. Three of the polysaccharides are common to more than one strain.     

Structural analysis of the sheath of a sheathed bacterium, Leptothrix cholodnii

Leptothrix cholodnii is an aerobic sheath-forming bacterium often found in oligotrophic and metal-rich aquatic environments. The sheath of this bacterium was isolated by selectively lysing the cells. Glycine and cysteine were the major amino acids of the sheath. The sheath was readily dissolved in hydrazine, and a polysaccharide substituted with cysteine was recovered from the solution. Galactosamine, glucosamine and galacturonic acid were detected in the hydrazinolysate by gas liquid chromatography analysis. FAB-MS analysis of the hydrazinolysate suggested a sugar sequence of HexN-GalA-HexN-HexN. Methylation linkage analysis revealed the presence of 4-linked GalA, 3-linked HexN and 4-linked HexN. The sulfhydryl groups of the sheath were used for labeling with the fluorogenic reagent, 4-(aminosulfonyl)-7-fluoro-2,1,3-benzoxadiazole (ABD-F). The labeled sheath (ABD-sheath) was partially hydrolyzed and three fluorescent fragments were purified by HPLC. One of them was identified as ABD-cysteine. The second one was found to be the ABD-cysteine tetramer. Another fragment was indicated to be a pentasaccharide substituted with ABD-cysteine by nuclear magnetic resonance (NMR) analysis. It can be assumed that the polysaccharide and peptide moieties of the sheath are connected by a cysteine residue. NMR analysis of the hydrazinolysate revealed that the polysaccharide moiety of the sheath was constructed from a pentasaccharide repeating unit containing 2-amino-2-deoxygalacturonic acid (GalNA), as shown below. -->4)-alpha-GalNA-(1-->4)-alpha-D-GalN(p)-(1-->4)-alpha-D-GalA(p)-(1-->4)-beta-D-GlcN(p)-(1-->3)-beta-D-GalN(p)-(1-->.     

Synthesis of non-immunosuppressive cyclophilin-Binding cyclosporin A derivatives as potential anti-HIV-1 drugs

Original cyclosporin A (CsA) derivatives bearing various alkylthio side chains at the sarcosine residue 3 (R configuration) and for the most potent and selective compounds a 4′-hydroxyl group at the Me-Leucine residue 4 were prepared in one or two steps from commercially available CsA. The [2-(dimethyl or diethylamino)-ethylthio-Sar]³-[(4′-OH)MeLeu]⁴-CsA derivatives 3k and 3l displayed potent in vitro anti-HIV-1 (IC₅₀ 46 nM) and low immunosuppressive activities (IC₅₀≥1500 nM).     

Cellouronate (β-1,4-linked polyglucuronate) lyase from Brevundimonas sp. SH203: Purification and characterization

Biodegradation of cellouronate (β-1,4-linked polyglucuronic acid sodium salt, β-1,4-linked glucuronan), which was prepared from regenerated cellulose by 2,2,6,6-tetramethylpiperidine-1-oxyl radical (TEMPO) mediated oxidation, was investigated. A bacterial strain with the ability to degrade cellouronate was isolated from soil collected in a natural environment, and identified as Brevundimonas sp. SH203 by comparing the nucleotide sequences of its 16S rDNA with those registered in the GenBank database. Cellouronate lyase-I (CUL-I), being responsible for the depolymerization of cellouronate, was purified to homogeneity from cell-free extracts. CUL-I was a monomeric protein with the molecular mass of 39 kDa by SDS–PAGE and 37 KDa by size exclusion chromatography (SEC). The enzyme activity was optimum at pH 7.5 and was inhibited by some divalent metal ions such as Mg²⁺, Fe²⁺ and Mn²⁺. The enzymatic reaction products were analyzed by SEC, TLC and ¹³C NMR. The results indicated that CUL-I catalyzed to depolymerize cellouronate endolytically to oligocellouronates and monomeric uronate.