Structural characterization of a pectic polysaccharide from Nerium indicum flowers.

A polysaccharide fraction, J6, was isolated from the hot-water extract of flowers of oleander Nerium indicum Mill., using ethanol precipitation, cetyltrimethylammonium bromide (CTAB) complexing, anion-exchange chromatography and gel permeation chromatography. J6 was found to contain L-rhamnose, L-arabinose, D-galactose, and D-galacturonic acid, in the ratio of 10.1:49.8:30.1:10.0. Its structure was investigated by methylation analysis, periodate oxidation, Smith degradation, partial acid hydrolysis, electrospray ionization mass spectrometry and NMR spectroscopic methods. It was found that J6 is an RG-I type polysaccharide, which contains a rhamnogalacturonan backbone, with various branches attached to O-4 of L-rhamnose. The branches probably involve (1-->4)-beta-D-galactan, branched L-arabino-(1-->3)(1-->6)-beta-D-galactan, and (1-->5)-alpha-L-arabinan. J6 stimulated NO production of macrophage RAW264.7 cells in a preliminary test.

Chemical structure and immunoinhibitory activity of a pectic polysaccharide containing glucuronic acid from the leaves of Diospyros kaki.

Pectin is a family of complex polysaccharides characterized by the presence of 1,4-linked alpha-galacturonic acid residues in the backbone and is more recognized for its potential bioactive properties. Herein we reported the structural and biological features of a pectic fraction DL-4OAC-1 from the leaves of Diospyros kaki. Chemical and spectral analysis revealed this polysaccharide possessed a backbone composed of a repeating disaccharide [-->4)-alpha-GalAp-(1-->2)-alpha-Rhap-(1-->], substituted at some O-4 of Rhap residues and O-3 of GalAp residues by the side chains consisting of Araf, Galp, Xylp, and in particular terminal GlcAp residues. The immunological assay in vitro showed that this distinct pectic fraction could inhibit the LPS-induced B lymphocyte proliferation and had no effect on the ConA-induced T lymphocyte proliferation.

Characterization of a pectic polysaccharide from the leaves of Diospyros kaki and its modulating activity on lymphocyte proliferation.

Pectin is a group of carbohydrate polymers constructing the primary cell walls and the middle lamella of terrestrial plants. Herein, we demonstrated the structure and immunomodulatory activity of the major pectic polysaccharide DL-3B(2) isolated from the leaves of Diospyros kaki. Based on composition analysis, methylation analysis, two-step acid hydrolysis, lithium-mediated selective degradation, (13)C NMR spectroscopy, and electrospray ionization mass spectrometry, DL-3B(2) was found to contain an alpha-1, 4-linked galacturonic acid (GalA) backbone with some insertions of alpha-1, 2-linked rhamnose residues. The arabinan- and arabinogalactan-side chains were attached to O-4 of the rhamnose residues, whereas the linear arabinoxylan was probably linked to O-3 of the GalA residues. Immunological tests in vitro showed that DL-3B(2) could help stimulate lipopolysaccharide-induced B lymphocyte proliferation, but not ConA-induced T lymphocyte proliferation, and that the arabinose residues play a role in maintaining this immunological activity.

New polysaccharide from Nerium indicum protects neurons via stress kinase signaling pathway.

Most of the polysaccharides purified from Chinese medicinal herbs showed anti-tumor and immune-stimulating effects. However, little is known about their effects on neuroprotection. Our previous study has demonstrated that polysaccharides (J2, J3 and J4) isolated from the flowers of Nerium indicum (Oleander) exert partial protection in cortical neurons stressed by beta-amyloid (Abeta) peptides or deprivation of nutrition from serum. In this study, we have isolated and characterized a new polysaccharide from the flowers of N. indicum (named as J6) and aimed to investigate its neuroprotective effects against Abeta-induced apoptosis. Pretreatment of the polysaccharide J6 significantly decreased the activity of caspase-3 as well as the cytotoxicity triggered by Abeta peptides in a dose-dependent manner. In contrast to the activation of survival signaling such as Akt found in J2, J3 and J4 fractions, neuroprotective effects of J6 markedly inhibited Abeta peptide-stimulated phosphorylation of c-Jun N-terminal kinase (JNK-1) as determined by Western blot analysis. Taken together, the polysaccharide J6 isolated from the flowers of N. indicum can serve as potential neuroprotective agent against neuronal death in Alzheimer's disease and the neuroprotective mechanism may primarily rely on inactivation of JNK signaling pathway.

Structural characterization and immunological activity of two cold-water extractable polysaccharides from Cistanche deserticola Y. C. Ma.

Two major polysaccharide fractions, CDA-1A and CDA-3B, were isolated from the cold-water extract of Cistanche deserticola Y. C. Ma, a holoparasitic plant and a valuable traditional Chinese medicine, using anion-exchange chromatography on DEAE-cellulose and gel-permeation chromatography on Sephacryl S-300 and Sephadex G-150. Their major structural features were elucidated using component and linkage analyses, periodate oxidation, Smith degradation, partial acid hydrolysis, and NMR spectroscopy. The results indicated that CDA-1A is an alpha-(1-->4)-D-glucan with alpha-(1-->6)-linked branches attached to the O-6 of branch points and that CDA-3B is an RG-I polysaccharide containing a typical rhamnogalacturonan backbone and arabinogalactan or arabinan branches. Bioactivity tests showed that CDA-1A is inert for T-cell proliferation stimulation but active for B-cell proliferation, while CDA-3B is potent for the stimulation of both T- and B-cell proliferation.

A beta-D-glucan isolated from the fruiting bodies of Hericium erinaceus and its aqueous conformation.

HEP3, a beta-D-glucan slightly soluble in water, was isolated from the alkaline extract of the fruiting bodies of Hericium erinaceus. Its chemical structure was investigated by methylation analysis, periodate oxidation, Smith degradation and by IR and NMR spectroscopy. It was shown to have a main chain composed of beta-(1-->3)-linked D-glucopyranosyl residues, with single unit glucosyl branches attached to O-6 of every third backbone residue. Viscometry and Congo red reaction indicated that HEP3 has a highly ordered hydrogen-bond dependent conformation in aqueous solution, which collapses in strong alkaline solution.

[Isolation, purification and structural analysis of a polysaccharide MDG-1 from Ophiopogon japonicus].

AIM: To separate and purify the anti-myocardial ischemic polysaccharide fraction with a homogenous molecular weight from Ophiopogon japonicus, then study the chemical structure of the parts. METHODS: Crude polysaccharides were prepared by extracting the tube root fraction of Ophiopogon japonicus with water, then precipitation with ethanol. From the crude polysaccharides, the polysaccharide of MDG-1 was separated and purified using ultrafiltration, DEAE Sepharose FF and Sephadex G-25 column chromatography. Its structure was studied by complete hydrolysis, periodate oxidation, Smith degradation, methylation analysis, 1H NMR and 13C NMR analysis etc. RESULTS AND CONCLUSION: MDG-1 was a water-soluble beta-D-fructosan, containing a backbone composed of Fruf (2 --> 1), and a branch of Fruf (2 --> 6) Fruf (2 --> per average 2. 8 of main chain residues. Mn, Mw and Mp of MDG-1 were 3 400, 4 800 and 5 000, respectively. MDG-1 contains trace of Glc, which maybe connect to its reducing terminal. Molar ratio of Fru and Glc is approximately 35: 1.

Structural investigation of a novel rhamnoglucogalactan isolated from the fruiting bodies of the fungus Hericium erinaceus.

A new heteropolysaccharide (HEP-1) was isolated from the fruiting bodies of Hericium erinaceus. It was estimated to have a molecular weight of 1.8x10(4) da and showed [alpha](D)(20) +129 (c 0.295, H(2)O). HEP-1 is composed of rhamnose, galactose, and glucose in the ratio of 1.19:3.81:1.00. Its structural features were investigated using composition analysis, methylation analysis, partial hydrolysis, and IR and NMR spectroscopy. The results showed that HEP-1 has a (1-->6)-linked alpha-d-galactopyranosyl backbone with branches that are composed of rhamnose and glucose attached to O-2.

Characterization of polysaccharides from the flowers of Nerium indicum and their neuroprotective effects.

Degeneration of neurons is a key problem in Alzheimer's disease (AD) and neuroprotection is a possible way to safeguard neurons from neurodegeneration. Polysaccharides isolated from Chinese medicinal herbs have been investigated extensively for their anti-tumor and immune stimulating effects. Yet, little is known about the effects of polysaccharides in neurons. Recently, two pure polysaccharides isolated from the flowers of Nerium indicum were shown to stimulate proliferation and differentiation of PC12 pheochromocytoma cells, an effect similar to that observed from nerve growth factor. In this notion, it is hypothesized that polysaccharides isolated from the flowers of N. indicum could exhibit beneficial effects in neurons. In this study, we isolated, characterized and investigated two new polysaccharides from the flowers of N. indicum for their neuroprotective effects on neurons against serum-deprivation and beta-amyloid (Abeta) peptide toxicity in primary rat cortical neuronal cultures. Pretreatment of the polysaccharides significantly reduced the number of apoptotic neurons revealed by DAPI staining when neurons were exposed to serum-free medium. Besides, the polysaccharides could also decrease the activity of caspase-3 triggered by Abeta peptides. Western blot analysis indicated that polysaccharides stimulated the phosphorylation of PDK-1 (Serine 241) and Akt (Threonine 308). In conclusion, the polysaccharides J2, J3 and J4 isolated from N. indicum provide a lead for future development of neuroprotective agent against neuronal death in neurodegenerative diseases and the neuroprotective mechanism may primarily rely on activation of Akt survival signaling pathway.

Structural analysis of a pectic polysaccharide from the leaves of Diospyros kaki.

A pectic polysaccharide DL-2A with a molar mass of 8.5 x 10(5), was obtained from the boiling water extract of Diospyros kaki leaves. It had [alpha]20D -21.8 degrees (c 0.22, H2O) and consisted of rhamnose, arabinose, galactose, xylose and galacturonic acid units in the molar ratio of 0.4:3.4:2.4:1.0:0.8, along with traces of glucuronic acid. About 16.7% of galacturonic acid existed as the methyl ester. A combination of linkage analyses, periodate oxidation, partial acid hydrolysis, selective lithium-degraded reaction, ESIMS, 1H- and 13C- NMR spectral analyses revealed its structural features. It was found that DL-2A possessed an alpha-(1-->4)-galacturonan backbone with some insertions of alpha-1,2-Rhap residues. The side-chains of arabino-3,6-galactan were attached to the backbone via O-4 of Rhap residues and O-3 of GalAp residues, while 4-linked xylose residues (forming short linear chains) were directly linked to O-4 of rhamnose residues, not as part of the xylogalacturonan. These novel structural features enlarge the knowledge on the fine structure of pectic substances in the plant kingdom.

Structure and potential immunological activity of a pectin from Centella asiatica (L.) Urban.

S3A was a RG-I pectin isolated from Centella asiatica that contained Rha, Ara, Gal, Glc and GalA in molar ratio of 1.0:0.6:1.5:0.2:1.1 and had been found to have a backbone composed mainly of the disaccharide repeat unit, -->4)-alpha-D-GalpA-(1-->2)-alpha-L-Rhap-(1-->. Based on methylation analysis, NaIO4 oxidation, partial acid hydrolysis and lithium-treatment, the structural features were elucidated. Side chains of S3A were predominantly linked to O-4 of 1,2,4-linked alpha-L-Rhap. The side chains are comprised of arabinosyl chains, galactosyl chains, arabinogalactosyl chains and short glucosyl chains. A total of 45% Rhap in the backbone was substituted by side chains. The arabinosyl residues were mostly distributed in the arabinosyl side chains. According to the immunological results of S3A and its degraded derivatives, S3A had no immunological activity, but its derivatives had immuno-stimulating activities to some extent.

Characterization of a rhamnogalacturonan and a xyloglucan from neriumindicum and their activities on PC12 pheochromocytoma cells.

Two polysaccharides, a rhamnogalacturonan (J1) and a xyloglucan (J2), were isolated and purified from the whole flowers of Nerium indicum and characterized by GC, GC-MS, NMR, and ESMS. The rhamnogalacturonan was found to consist of repeating units of a linear trisaccharide with the structure [-->4)-alpha-GalpA(1-->2)alpha-Rhap(1-->4)-alpha-GalpA(1-->4)-](n)(), while the xyloglucan consisted of repeating units of the trisaccharide, which was (1-->4)-linked beta-glucopyranosyl residues as a backbone and substituted at position 6 linked to terminal alpha-xylopyranose side chains. These two polysaccharides were tested on the proliferation and differentiation on PC12 pheochromocytoma cells and found to have effects similar to nerve growth factor (NGF).

Structural characterization of a water-soluble beta-D-glucan from fruiting bodies of Agaricus blazei Murr.

A beta-D-glucan, Ab2-2N, was isolated from the hot-water extract of fruiting bodies of Agaricus blazei Murr by ethanol precipitation, anion-exchange and gel-permeation chromatography. Its structure was investigated by composition analysis, methylation analysis, Smith degradation, mild hydrolysis, and NMR spectroscopy. It contains a (1-->6)-linked beta-D-glucopyranosyl backbone, with one side chain consisting of terminal and 3-substituted beta-D-glucopyranosyl residues attached at O-3 for every three backbone residues.

Purification, characterization, and modification of T lymphocyte-stimulating polysaccharide from spores of Ganoderma lucidum.

The hot-water extract of the spores of Ganoderma lucidum was shown to have a stimulating effect on concanavalin A-induced mitogenic activity of T lymphocytes. Bioassay-guided separation led to the isolation of a polysaccharide with potent T lymphocyte-stimulating activity by ethanol fractionation, anion-exchange, and size-exclusion chromatography. Based on the composition and methylation analyses, periodate oxidation, Smith degradation, and NMR spectroscopy, the native polysaccharide was shown to be a beta-D-(1-->3)-glucan with branches of terminal glucosyl residues substituted at C-6 of the glucose residues in the main chain. The branching ratio is approximately 20%. A series of sulfated or carboxymethylated derivatives were prepared and their structural features were elucidated by chemical and spectral analyses. The solution conformation and T lymphocyte proliferation effect of the glucans before and after derivatization were compared and discussed. The data obtained indicate that the introduction of ionic groups would significantly affect the original conformation of the native glucan in aqueous solution and further affect T lymphocyte-stimulating activity. The triple-helical structure of the glucans, the nature of the ionic groups, and the density of negative charge were considered to be closely related to this activity.

Structural features of an immunostimulating and antioxidant acidic polysaccharide from the seeds of Cuscuta chinensis.

Three crude polysaccharide fractions, named CS-A, CS-B and CS-C, were prepared from the seeds of Cuscuta chinensis by hot water extraction and diluted alkali extraction subsequently, then EtOH precipitation, and tested for lymphocyte proliferation activity. CS-A showed a stimulating effect on concanavalin A or lipopolysaccharide induced mitogenic activity of lymphocytes. An acidic polysaccharide (CS-A-3beta) was purified from CS-A by anion exchange and gel filtration chromatography. The polysaccharide showed potent stimulating effects on lymphocyte proliferation and antibody production, but did not significantly influence the serum IgG level. Its structural features were elucidated by methylation analysis, partial acid hydrolysis, 1D and 2D NMR and ESI-mass spectroscopy. The data obtained indicated that CS-A-3beta had a backbone consisting of alpha-D-1,4-linked GalpA residues and beta-L-1,2-linked Rhap residues with branches at C-4 of Rhap residues and C-3 of GalpA residues, composed of arabinogalactan and glucobiose. This structure is typical for a pectic polysaccharide of the rhamnogalacturonan type. In addition, the effect of CS-A, CS-B, CS-C and CS-A-3beta on hydrogen peroxide induced cell lesion in rat pheochromocytoma line PC 12 was investigated. The results indicated that, besides its immunostimulating activity, CS-A-3beta had a protective effect against free radical-induced cell toxicity.

Structural characterization and biological activities of SC4, an acidic polysaccharide from Salvia chinensis.

AIM: To study the chemical characterization and some biological activities of an acidic polysaccharide, named SC4 from Salvia chinensis. METHODS: Chemical and spectral methods were employed to identify the structural characterization of SC4. Effect of SC4 on the proliferation of T- and B-lymphocytes both in vivo and in vitro was measured at various concentrations, and the protective effects of SC4 on PC12 cells against H2O2-induced injury was observed. RESULTS: SC4 was a highly branched polysaccharide with mean molecular weight of 4.5 10(5), composed of Rha, Xyl, Ser, Gal, and GalA in the molar ratio of 1.0 7.0 5.3 1.2 4.2. Methylation analysis and NMR identified the linkages of the residues of SC4. SC4 increased the B-lymphocyte proliferation and spleen weight remarkably while no obvious changes were examined in T-lymphocyte proliferation, thymus weight, and the level of IgG and C3 in blood of mice. Moreover, SC4 enhanced the PC12 cells viability after H2O2 treatment. CONCLUSION: SC4, the acidic polysaccharide with complicated structure, was a B-lymphocyte stimulator and protected PC12 cells at the concentration of 20 mg/L against H2O2-induced injury.

Structural features of immunologically active polysaccharides from Ganoderma lucidum.

Three polysaccharides, two heteroglycans (PL-1 and PL-4) and one glucan (PL-3), were solubilized from the fruit bodies of Ganoderma lucidum and isolated by anion-exchange and gel-filtration chromatography. Their structural features were elucidated by glycosyl residue and glycosyl linkage composition analyses, partial acid hydrolysis, acetolysis, periodate oxidation, 1D and 2D NMR spectroscopy, and ESI-MS experiments. The data obtained indicated that PL-1 had a backbone consisting of 1,4-linked alpha-D-glucopyranosyl residues and 1,6-linked beta-D-galactopyranosyl residues with branches at O-6 of glucose residues and O-2 of galactose residues, composed of terminal glucose, 1,6-linked glucosyl residues and terminal rhamnose. PL-3 was a highly branched glucan composed of 1,3-linked beta-D-glucopyranosyl residues substituted at O-6 with 1,6-linked glucosyl residues. PL-4 was comprised of 1,3-, 1,4-, 1,6-linked beta-D-glucopyranosyl residues and 1,6-linked beta-D-mannopyranosyl residues. These polysaccharides enhanced the proliferation of T- and B-lymphocytes in vitro to varying contents and PL-1 exhibited an immune-stimulating activity in mice.

[Structural features of a neutral heteropolysaccharide CPB-4 from Cynanchum paniculatum].

OBJECTIVE: To study the chemical features of CPB-4, a heteropolysaccharide obtained from Cynanchum paniculatum. METHOD: Sugar composition analysis, methylation analysis, partial hydrolysis and carbon-13 nuclear magnetic resonance were used to determine the sugar composition, linkages, main chain, branch chains and branching points. RESULT: CPB-4 is composed of L-arabinose, L-xylose, L-rhamnose and D-galactose in closely molar ratios of 0.8:0.2:0.2:1.0. Its main chain is comprised of 1, 5 linked galactose and side chains are comprised of terminal xylose, terminal arabinose, oligosaccharide of arabinose and oligosaccharide of arabinose, rhamnose and galactose. The branching points are located at C-6 and C-2 of galactose. CONCLUSION: CPB-4 is a new heteropolysaccharide from C. paniculatum.

[Chemical studies on SC3, a polysaccharide from Salvia chinensis].

AIM: To study the chemical structure of SC3, an acidic polysaccharide from Salvia chinesis. METHODS: Based on chemical (including sugar composition analysis, methylation analysis, uronic acid reduction and partial acid hydrolysis) and spectral analysis (IR, NMR, ESI-MS), the structural characterization of SC3 was investigated. RESULTS: SC3 was composed Rha, Ara, Gal and GalA, with its mean molecular weight of 7.7 x 10(4). By means of methylation analysis, partial acid hydrolysis, NMR and ESI-MS spectrum, the linkages and sequence information of SC3 were obtained. CONCLUSION: SC3 is an complicated acidic polysaccharide, obtained for the first time from the plant.