A new naturally acetylated triterpene saponin from Nigella sativa

A new glycosylated triterpene 1 was identified as 3-O-[β-d-xylopyranosyl-(1→3)-α-l-rhamnopyranosyl-(1→4)-β-d-glucopyranosyl]-11-methoxy-16-hydroxy-17-acetoxy hederagenin from an ethanolic extract of seeds of Nigella sativa Linn. Identification of the naturally acetylated saponin was based on chemical and spectroscopic analyses including FABMS, ¹H, ¹³C, and 2D NMR and DEPT. The saponin was a penta hydroxy pentacyclic triterpene, in which one hydroxyl group was acetylated and other one was methylated naturally.     

Structural characterisation of novel lichen heteroglycans by NMR spectroscopy and methylation analysis

Two galactofuranomannans, Ths-4 and Ths-5, were isolated from the lichen, Thamnolia vermicularis var. subuliformis, using ethanol fractionation and anion-exchange and size-exclusion chromatography. The average molecular weights of Ths-4 and Ths-5 were estimated to be 19 and 200 kDa, respectively. Structural characterisation of Ths-4, Ths-5 and their partially hydrolysed derivatives was performed by methanolysis and methylation analysis. The intact and partially hydrolysed Ths-4 was further analysed using NMR spectroscopy (1D, COSY, NOESY, TOCSY, HSQC and HMBC). According to the data obtained, the heteroglycans Ths-4 and Ths-5 have similar structures, but have large differences in molecular weight. The structure is composed of 3-O-linked and 5-O-linked galactofuranosyl chains linked to a mannan core. The mannan core consists of a main chain of alpha-(1-->6)-linked mannopyranosyl residues, substituted at O-2 with either a single alpha-mannopyranosyl unit or an alpha-Manp-(1-->2)-alpha-Manp-(1-->2)-alpha-Manp group in the ratio of approximately 1:3, respectively. The polysaccharides have idealised repeating blocks as is shown.     

Structural investigations on betacyanin pigments by LC NMR and 2D NMR spectroscopy

Four betacyanin pigments were analysed by LC NMR and subjected to extensive NMR characterisation after isolation. Previously, low pH values were applied for NMR investigations of betalains resulting in rapid degradation of the purified substances thus preventing extensive NMR studies. Consequently, up to now only one single (13)C NMR spectrum of a betalain pigment, namely that of neobetanin (=14,15-dehydrobetanin), was available. Because of its sufficient stability under highly acidic conditions otherwise detrimental for betacyanins, this pigment remained an exemption. Since betalains are most stable in the pH range of 5-7, a new solvent system has been developed allowing improved data acquisition through improved pigment stability at near neutral pH. Thus, not only (1)H, but for the first time also partial (13)C data of betanin, isobetanin, phyllocactin and hylocerenin isolated from red-purple pitaya [Hylocereus polyrhizus (Weber) Britton & Rose, Cactaceae] could be indirectly obtained by gHSQC- and gHMQC-NMR experiments.     

Structural elucidation of a 3-O-methyl-D-galactose-containing neutral polysaccharide from the fruiting bodies of Phellinus igniarius

PIP60-1, a novel heteropolysaccharide isolated from fruiting bodies of the medicinal fungus, Phellinus igniarius, has a molecular weight of 1.71 x 10(4)Da and is composed of L-fucose, D-glucose, D-mannose, D-galactose and 3-O-Me-D-galactose in a ratio of 1:1:1:2:1. A structural investigation of PIP60-1 carried out using sugar and methylation analyses, combined with (1)H and (13)C NMR spectroscopy, including COSY, TOCSY, NOESY, HSQC and HMBC experiments, established the repeating unit of the polysaccharide as the following: [structure: see text]     

Structural elucidation of a cell wall fungal polysaccharide isolated from Ustilaginoidea virens, a pathogenic fungus of Oriza sativa and Zea mays

The alkali-extractable water-soluble polysaccharides (F1SS) isolated from the outer cell wall of two strains of Ustilaginoidea virens have been studied by chemical and methylation analyses, and 1D and 2D ¹H and ¹³C NMR spectroscopy. The structures of these polysaccharides are very similar, and can be described by the following idealized repeating unit: where n and m are approximately 1 and 2, respectively.     

Novel acetylated alpha-cyclosophorotridecaose produced by Ralstonia solanacearum

Alpha-cyclosophorotridecaose (alpha-C13) produced by Ralstonia solanacearum is isolated by trichloroacetic acid treatment and subjected to various chromatographic techniques. Here, we report for the first time that R. solanacearum produces acetylated alpha-C13. Structural analyses of the acetylated alpha-C13 were performed with 1D or 2D NMR spectroscopy, MALDI-TOF MS and HPLC. The results show that the alpha-C13 is substituted by mainly one acetyl residue at the C-6 position of the glucose unit.     

Characterization of an acetylated heteroxylan from Eucalyptus globulus Labill

A heteroxylan was isolated from Eucalyptus globulus wood by extraction of peracetic acid delignified holocellulose with dimethyl sulfoxide. Besides (1-->4)-linked beta-D-xylopyranosyl units of the backbone and short side chains of terminal (1-->2)-linked 4-O-methyl-alpha-D-glucuronosyl residues (MeGlcA) in a 1:10 molar ratio, this hemicellulose contained galactosyl and glucosyl units attached at O-2 of MeGlcA originating from rhamnoarabinogalactan and glucan backbones, respectively. About 30% of MeGlcA units were branched at O-2. The O-acetyl-(4-O-methylglucurono)xylan showed an acetylation degree of 0.61, as determined by 1H NMR spectroscopy, and a weight-average molecular weight (M(w)) of about 36 kDa (P=1.05) as revealed from size-exclusion chromatography (SEC) analysis. About half of the beta-D-xylopyranosyl units of the backbone were found as acetylated moieties at O-3 (34 mol%), O-2 (15 mol%) or O-2,3 (6 mol%). Practically, all beta-D-xylopyranosyl units linked at O-2 with MeGlcA residues were 3-O-acetylated (10 mol%).     

Evidence for cross-linking in tomato cutin using HR-MAS NMR spectroscopy

Cutin is a polyester biopolymer component of plant leaf and fruit cuticles, most often associated with waxes and cuticular polysaccharides, and sometimes with another aliphatic biopolymer called cutan. Insolubility of these cuticular biopolymers has made it difficult to apply traditional analytical techniques for structure determination, because most techniques providing molecular level details require solubility. By using the relatively new technique of one and two-dimensional high-resolution magic angle spinning (HR-MAS) NMR spectroscopy, with added information from solid-state 13C NMR spectroscopy, detailed through-bond connectivities and assignments are made for cutin from Lycopersicon esculentum (tomato) fruit. Based on the data obtained, tomato cutin is found to be predominantly an aliphatic polyester with some olefinic and aromatic moieties, consistent with previous studies that employed various degradative approaches. Aside from esters, there are free primary and secondary alcohol groups, as well as free fatty acids. A significant finding is the presence of alpha-branched fatty acids/esters. Mid-chain hydroxyls appear to be generally unesterified, but esters of mid-chain hydroxyls have been identified. The alpha-branched fatty acids/esters and esters of mid-chain hydroxyls could point towards cross-linking.     

Refinement of the structures of cell-wall glucans of Schizosaccharomyces pombe by chemical modification and NMR spectroscopy

Alkali extraction and methylation analyses in the 1970s revealed that the cell walls of the yeast Schizosaccharomyces pombe contain a (1-->3)-alpha-d-glucan, a (1-->3)-beta-d-glucan, a (1-->6)-beta-d-glucan, and a alpha-galactomannan. To refine the structures of these polysaccharides, cell-wall glucans of S. pombe were extracted, fractionated, and analyzed by NMR spectroscopy. S. pombe cells were treated with 3% NaOH, and alkali-soluble and insoluble fractions were prepared. The alkali-insoluble fraction was treated with 0.5M acetic acid or Zymolyase 100T to yield an alkali-insoluble, acetic acid-insoluble fraction, an alkali-insoluble, Zymolyase-insoluble fraction, and an alkali-insoluble, Zymolyase-soluble fraction. (13)C NMR and 2D-NMR spectra disclosed that the cell wall of S. pombe is composed of three types of glucans, specifically, a (1-->3)-alpha-d-glucan, a (1-->3)-beta-d-glucan, which may either be linear or slightly branched, and a highly branched (1-->6)-beta-d-glucan, in addition to alpha-galactomannan. The highly branched (1-->6)-beta-d-glucan was identified by selective periodate degradation of side-chain glucose as a highly (1-->3)-beta-branched (1-->6)-beta-d-glucan with more branches than that of Saccharomyces cerevisiae. Flexibility of these polysaccharides in the cell wall was analyzed by (13)C NMR spectra in D(2)O. The data collectively indicate that (1-->3)-alpha- and (1-->3)-beta-d-glucans are rigid and contribute to the cell shape, while the highly branched (1-->6)-beta-d-glucan and alpha-galactomannan are flexible.     

Synthesized oversulfated and acetylated derivatives of polysaccharide extracted from Enteromorpha linza and their potential antioxidant activity

Sulfated polysaccharide extracted from blue algae Enteromorpha linza is proved to possess excellent antioxidant activities. Two derivatives by means of oversulfated and acetylated coupling were synthesized and investigated their antioxidant activities including scavenging effect of superoxide, hydroxyl and 1,1-diphenyl-2-picrylhydrazyl radicals. And then the relationship between antioxidant activity and chemical characteristics was characterized. The results of chemical analysis and FT-IR showed that the modification of polysaccharide was successful. In addition, it was found that certain derivatives exhibited stronger antioxidant activity than raw material. They could serve as free-radical inhibitors or scavengers, acting possibly as primary antioxidants.     

Structural elucidation of the polysaccharide moiety of a glycopeptide (GLPCW-II) from Ganoderma lucidum fruiting bodies

A water-soluble glycopeptide (GLPCW-II) was isolated from the fruiting bodies of Ganoderma lucidum by DEAE-Sepharose Fast-Flow and Sephacryl S-300 High Resolution Chromatography. The glycopeptide had a molecular weight of 1.2x10(4)Da (determined by HPLC), and consisted of approximately 90% carbohydrate and approximately 8% protein as determined using the phenol-sulfuric acid method and the BCA protein assay reagent kit, respectively. The polysaccharide moiety was composed mainly of D-Glc, L-Fuc, and D-Gal in the ratio of 1.00:1.09:4.09. To facilitate structure-activity studies, the structure of the GLPCW-II polysaccharide moiety was elucidated using 1H and 13C NMR spectroscopy including COSY, TOCSY, HMBC, HSQC, and ROESY, combined with GC-MS of methylated derivatives, and shown to consist of repeating units with the following structure: [Formula: see text].     

Structural characterization of the acetylated heteroxylan from the natural hybrid Paulownia elongata/Paulownia fortunei

The heteroxylan from the hybrid Paulownia elongata/Paulownia fortunei is an O-acetyl-(4-O-methylglucurono)xylan with an acetylation degree (DS) of 0.59 and a molecular weight (M(w)) of 29 kDa. The heteroxylan backbone is composed by (1-->4)-linked beta-d-xylopyranosyl units (Xylp) partially ramified with terminal (1-->2)-linked 4-O-methyl-alpha-D-glucuronosyl (MeGlcpA) and a small proportion of alpha-D-glucuronosyl (GlcpA) residues in a molar ratio of Xylp:(MeGlcpA+GlcpA) of 20:1. Roughly half of the beta-D-xylopyranosyl units in the backbone are acetylated: 3-O-acetylated (22 mol %), 2-O-acetylated (23 mol %) or 2,3-di-O-acetylated (7 mol %). ESI-MS and MALDI-MS studies of partially hydrolyzed heteroxylan revealed a random distribution of O-Ac and MeGlcpA within the backbone. However, the frequency of substitution with O-Ac along the backbone is not uniform and the molecular regions that did not contain MeGlcpA substituents possessed an acetylation degree significantly lower than the average DS of the xylan.     

Structural elucidation of the core-lipid A backbone from the lipopolysaccharide of Acinetobacter radioresistens S13, an organic solvent tolerant Gram-negative bacterium

The structure of the core oligosaccharide of the lipopolysaccharide from an organic solvent tolerant Gram-negative bacterium, Acinetobacter radioresistens S13, was investigated by chemical analysis, NMR spectroscopy and MALDI-TOF mass spectrometry. All the experiments were performed on the oligosaccharides obtained either by alkaline degradation or mild acid hydrolysis. The data showed the presence of two novel oligosaccharide molecules containing a trisaccharide of 3-deoxy-D-manno-octulopyranosonic acid in the inner core region and a glucose rich outer core whose structure is the following: [structure: see text] R=H in the main oligosaccharide and beta-Glc in the minor product. The bacterium was grown on aromatic (phenol and benzoic acid) and nonaromatic carbon sources and the core oligosaccharide resulted to occur always with this novel structure.     

Metabolomic analysis of the consequences of cadmium exposure in Silene cucubalus cell cultures via 1H NMR spectroscopy and chemometrics

Several essential and non-essential metals (typically those from periods 4, 5 and 6 in groups 11-15 in the periodic table) are commonly detoxified in higher plants by complexation with phytochelatin. The genetic and gross metabolic basis of metal tolerance in plants is, however, poorly understood. Here, we have analyzed plant cell extracts using 1H NMR spectroscopy combined with multivariate statistical analysis of the data to investigate the biochemical consequences of Cd(2+) exposure in Silene cucubalus cell cultures. Principal components analysis of 1H NMR spectra showed clear discrimination between control and Cd(2+) dosed groups, demonstrating the metabolic effects of Cd(2+) and thus allowing the identification of increases in malic acid and acetate, and decreases in glutamine and branched chain amino acids as consequences of Cd(2+) exposure. This work shows the value of NMR-based metabolomic approaches to the determination of biochemical effects of pollutants in naturally selected populations.     

An acetylated galactoglucomannan from Picea abies L. Karst

A water-soluble galactoglucomannan composed of D-galactose, D-glucose, and D-mannose in 1:3:17 mole proportion has been isolated from the secondary cell walls of Picea abies L. Karst. About 33% of the polysaccharide units were substituted by acetyl groups. Structural studies of the polymer indicated a beta-(1-->4)-linked glucomannopyranosyl backbone with a low content of branch points at O-6 of mannosyl and glucosyl residues. A preference for mannosyl groups indicates the presence of a single D-galactosyl unit side-chain. About half of the mannose residues were O-acetylated at C-2 and C-3 in 1.7:1 mole proportion.     

Structure of a major glycolipid from Thermus oshimai NTU-063

The structure of a major glycolipid isolated from the thermophilic bacteria Thermus oshimai NTU-063 was elucidated. The sugar and fatty acid compositions were determined by GC-MS and HPLC analysis on their methanolysis and methylation derivatives, respectively. After removal of both O- and N-acyl groups by alkaline treatment, the glycolipid was converted to a fully acetylated tetraglycosyl glycerol derivative, the structure of which was then determined by NMR spectroscopy (TOCSY, HSQC, HMBC). Thus, the complete structure of the major glycolipid from T. oshimai NTU-063 was established as beta-Glcp-(1-->6)-beta-Glcp-(1-->6)-beta-GlcpNAcyl-(1-->2)-alpha-Glcp-(1-->1)-glycerol diester. The N-acyl groups on the 2-amino-2-deoxy-glucopyranose residue are C15:0 and C17:0 fatty acids, whereas the fatty acids of glycerol diester are more heterogeneous including both straight and branched fatty acids from C15:0 to C18:0.     

Structural elucidation of fungal polysaccharides isolated from the cell wall of Plectosphaerella cucumerina and Verticillium spp

The structure of acidic fungal polysaccharides isolated from the cell wall of Plectosphaerella cucumerina, Verticillium dahliae, and V. albo-atrum has been investigated by chemical analysis, methylation analysis, and 1D and 2D 1H and 13C NMR spectroscopy. The polysaccharides have an idealized repeating block of the type: [carbohydrates: see text] linked to a small mannan core (<15%), where n=13, m=13, p=5, and q=8 for P. cucumerina, and n=16, m=16, p=6, and q <1 for both Verticillium species.     

Wound-induced green leaf volatiles cause the release of acetylated derivatives and a terpenoid in maize

Green leaf volatiles (GLVs), generally occurring C6 alcohols, aldehydes and acetates from plants, play an important role in plant-plant communication. These compounds induce intact plants to produce jasmonic acid, and induce defense-related gene expression and the release of volatile compounds. Here, we address wound-induced GLVs cause the release of acetylated derivatives and a terpenoid, (E)-4,8-dimethylnona-1,3,7-triene (DMNT) in intact maize, which may be a type of plant-plant interaction mediated by airborne GLVs. Upon exposure of intact maize seedlings to wound-induced GLVs, (Z)-3-hexenyl acetate was consistently the most abundant compound released. Exogenous application of individual alcohols and aldehydes mostly resulted in the release of corresponding acetate esters. C6-alcohols with a double bond between the second and third, or the third and fourth carbon atoms, C5- or C6-aldehydes, and (Z)-3-hexenyl acetate triggered the release of DMNT. When (Z)-3-hexenyl acetate and hexyl acetate were used to treat maize seedlings, they were recovered from the plants. These data demonstrated that: (1) apart from direct adsorption and re-release of acetate esters, absorption and conversion of exogenous alcohols and aldehydes into acetate esters occurred, and (2) DMNT was induced by a range of aldehydes and unsaturated alcohols.     

NMR and MALDI-TOFMS analysis of a heteroglycan isolated from hot water extract of edible mushroom, Volvariella bombycina

A water-soluble polysaccharide, isolated from the hot aqueous extract of an edible mushroom, Volvariella bombycina, consists of d-glucose, d-mannose, and d-galactose in a molar ratio 2:1:1. On the basis of total hydrolysis, methylation analysis, periodate oxidation, and NMR studies ((1)H, (13)C, TOCSY, DQF-COSY, NOESY, ROESY, HMQC, and HMBC), and MALDI-TOFMS analysis, the repeating unit of the polysaccharide is established as.