Cold Water-soluble Polysaccharides Research Articles

Penthorum chinense Pursh polysaccharide induces a mitochondrial-dependent apoptosis of H22 cells and activation of immunoregulation in H22 tumor-bearing mice

Penthorum chinense Pursh has been widely used as a Chinese traditional functional food for liver disease prevention and therapy. A novel cold-water soluble polysaccharide (PCP-4) has been prepared from Penthorum chinense Pursh at 4 °C. PCP-4 contained both α and β configurations and mainly consisted of galactose, arabinose, galacturonic acid, rhamnose, and glucose (molar ratio: 6.38:5.26:1.69:1:0.87) with an average molecular weight of 1.83 × 106 Da. In vitro experiments results demonstrated that PCP-4 effectively inhibits the proliferation of H22 cells and blocks cells at S phase in a dose-dependent manner. Otherwise, PCP-4-treated cells exhibited typical morphological features of apoptotic cells. Combined the results from western blot analysis, Annexin V-FITC/PI staining, Rhodamine 123 staining and DCFH-DA staining, PCP-4 was found to induce H22 cell apoptosis in vitro via the mitochondrial signaling pathway. The results of in vivo experiments suggested that PCP-4 significantly suppresses xenograft tumor growth by protecting immune organs, enhancing immune cells functionality, and inducing apoptosis. Overall, these results clearly show that Penthorum chinense Pursh polysaccharide serves as a valuable natural product for hepatocellular carcinoma therapy.

A heteropolysaccharide from Rhodiola rosea L.: preparation, purification and anti-tumor activities in H22-bearing mice

Numerous polysaccharides isolated from plants have been used to augment traditional drugs in the treatment of cancer. In order to explore the influence to hepatocellular carcinoma, a novel cold water-soluble polysaccharide was separated from Rhodiola rosea L. root (RLP) and then its structure and anti-cancer activities were tested. The chemical compositions and high performance gel permeation chromatography (HPGPC) results indicated that RLP was an acid heteropolysaccharide with the molecular weight of about 1.15×106 Da. Furthermore, ion chromatography (IC), Fourier transform infrared (FT-IR) and nuclear magnetic resoance (NMR) further indicated that RLP was main composed of →2,4)-α-Rha(1→, →5)-α-L-Araf-(1→, α-D-Glu, →6)-β-D-Galp-(1→, β-D-Man and →4)-α-GalpA-(1→. In vivo antitumor activities of RLP were carried out by using H22 tumor-bearing mice model. The results shown that RLP (100 and 300 mg/kg) could inhibit tumor growth of H22 cells from 23.59 % to 45.52 % and protect thymuses and spleen without damage. In addition, according to cell cycle, AV-FITC/PI and JC-1, RLP could induce dose-dependent apoptosis of H22 cells via S phase arrested which was through a mitochondrial related pathway. Our data indicated that RLP has a broader application prospect in anti-tumor preparations.

The Structural Characterization of a Novel Water-Soluble Polysaccharide from Edible Mushroom Leucopaxillus giganteus and Its Antitumor Activity on H22 Tumor-Bearing Mice.

In the present study, a novel cold water-soluble polysaccharide fraction (LGP) with the average molecular weight of 1.78×106 Da was extracted and purified from Leucopaxillus giganteus and its primary structure as well as in vivo antitumor activity was evaluated. The monosaccharide composition of LGP was determined by ion chromatography to be galactose, xylose, glucose and fucose in a molar ratio of 2.568 : 1.209 : 1 : 0.853. Its backbone was composed of α-D-Glu, α-D-Xyl, α-D-Gal and α-L-Fuc. The results of in vivo antitumor experiment demonstrated that LGP could effectively protect immune organs, has excellent antitumor activity, and inhibit the proliferation of H22 solid tumors in a dose-dependent manner. By analyzing Annexin V-FITC/PI staining, cell cycle and mitochondrial membrane potential detection assay, we concluded that LGP induced apoptosis of H22 cells via S phase arrest and mitochondria-mediated apoptotic pathway. Our results could provide valuable information for the potential application of LGP as an anti-hepatoma agent.

A unique polysaccharide containing 3-O-methylarabinose and 3-O-methylgalactose from Tinospora sinensis

Tinospora sinensis (Lour.) Merrill is of great therapeutic significance in Indian traditional medicine. Crude polysaccharides were isolated from methanol pre-extracted stems of dried material by successive extractions with cold water, hot water and NaOH (0.25 mol/L) in 0.98, 0.55 and 0.70 % yields respectively. Cold water soluble polysaccharides (CWSP) were purified and fractionated by ion exchange chromatography on DEAE-Sephacel. Neutral polysaccharides were further fractionated on Sepharose CL6B to yield three fractions TW1, TW2, TW3. The study further focuses on structural elucidation of TW1. TW1 was obtained in 0.8 % yield relative to CWSP, with MW of 1.6 × 105 Da. It was composed of 3-O-methyl-arabinose, 3-O-methyl-galactose and galactose in molar ratio of 1.0:6.3:0.9 respectively. Based on per-deuteromethylation, NMR and ESI–MS analyses, TW1 was composed of 1,4-linked 3-O-methyl-β-d-galactopyranose and β-d-galactopyranose backbone with branching at O-6 of 3-O-methyl-β-d-galactosyl residues by 1,5-linked 3-O-methyl-α-l-arabinofuranoside chains. 3-O-methyl-arabinose and 3-O-methyl-galactose have first ever been reported in any polysaccharide and Tinospora genus, respectively.

Antrodia cinnamomea Galactomannan Elicits Immuno-stimulatory Activity Through Toll-like Receptor 4.

Antrodia cinnamomea (A. cinnamomea) is a medicinal fungus used in traditional Chinese medicine to treat different kinds of ailments, including liver diseases, abdominal pain, drug intoxication, diarrhea, itchy skin, hypertension, and cancer. Polysaccharides have been identified as one of the major pharmacologically active ingredients present in A. cinnamomea. The present study aims to investigate the immunoenhancing activity of galactomannan isolated from A. cinnamomea. The cold water-soluble polysaccharide (galactomannan-repeated; MW>70 kDa; named ACP) of A. cinnamomea was isolated, and immunostimulatory properties were studied through different immune cell models including mouse macrophages and human dendritic cells. Through Toll-like receptor 4, ACP stimulated tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6) in J774A.1 mouse macrophages, mouse peritoneal macrophages and human dendritic cells. It was further identified that ACP elicited its immunostimulatory activity through protein kinase C-α (PKC-α) and mitogen activated protein kinases (MAPK) phosphorylation. Furthermore, ACP exerted the endotoxin tolerance-like effect through NF-κB inhibition. These findings demonstrate the potential of A. cinnamomea galactomannan as an immunostimulator or an adjuvant in immunotherapy and vaccination.

Acidic and Neutral Monosaccharide Analysis of Cold Water Soluble Polysaccharide from Hippophae salicifolia D. Don Leaves

Hippophae salicifolia D. Don is a shrub used in traditional medicine system since ages due to its diverse pharmacological effects and health benefits. The leaves of the species have immense therapeutic applications viz. antioxidant, antibacterial, anti-microbial, anti-inflammatory, immuno-modulatory etc. due to diverse array of chemical constituents. The cold water soluble polysaccharide (CWSP) was isolated from the leaves in 3.6% yield. Complete hydrolysis of the polysaccharide was accomplished using 2N sulphuric acid. Paper chromatography and gas liquid chromatography (GLC) of the hydrolysate identified that CWSP was composed of L-rhamnose, L-arabinose, D-xylose, D-mannose, D-galactose and D-glucose in the mole percent of 13.2, 21.5, 4.2, 9.4, 29.3 and 22.4 respectively. Uronic acid estimation was performed by reduction of the acidic moiety in polysaccharide followed by derivatisation to alditol acetates and GLC analysis of resultant neutral polysaccharide which led to quantification of D-galacturonic acid in 9.6 mole percent. The study resulted in composition of constituent monosaccharides of CWSP alongwith uronic acid content. The results may be useful in determination of structural moiety of the polysaccharide responsible for its bio-efficacy.

Do assorted approaches aid in estimation of uronic acids? Case studies on Tinospora sinensis polysaccharides

Polysaccharides containing uronic acids are of immense biological significance. Colorimetric methods using different chromogens have been explored for uronic acid estimation in polysaccharides but these methods counter numerous difficulties, when neutral sugars are present in substantial amount. The uronic acid content of cold water-soluble polysaccharide (CWSP) and hot water-soluble polysaccharide (HWSP) isolated from Tinospora sinensis, widely known for marked therapeutic significance has been studied. The uronic acid was quantitatively estimated by spectrophotometric methods using carbazole, 3-phenylphenol and 3,5-dimethylphenol (DMP) vis-a-vis gas–liquid chromatography (GLC) analysis. Galacturonic acid in CWSP and HWSP was estimated as 1.41% and 1.20% respectively. The findings bring to light, the efficacy of chromogens and also reveal to what extent, they could be relied upon. The results also indicated higher specificity and selectivity of DMP over carbazole and 3-phenylphenol. However, GLC analysis despite time consuming remains the finest method for precise estimation of uronic acids. Further, GLC analysis of alditol acetates indicated the monosugar composition of l-rhamnose, l-arabinose, d-mannose, d-galactose and d-glucose in molar ratio 0.178, 0.259, 0.140, 1.045 and 98.377 in CWSP and l-rhamnose, l-arabinose, d-xylose, d-mannose, d-galactose and d-glucose in molar ratio 0.608, 0.628, 0.978, 0.227, 0.111 and 97.448 in HWSP respectively.

Identification of Two Cold Water-Soluble Polysaccharides from the Stems of Ephedra sinica Stapf

Two polysaccharides (ESP-A1 and ESP-A2) were isolated from the cold water extract of Ephedra sinica Stapf and purified through ethanol precipitation, deproteinization and by ion exchange and gel-filtration chromatography. Their molecular weight was determined using high performance size exclusion chromatog-raphy and evaporative light scattering detector (HPSEC-ELSD) and their monosaccharide composition was analyzed by high performance capillary electrophoresis (HPCE) based on pre-column derivatization with 1-phenyl-3-methyl-5-pyrazolone (PMP). It was shown that ESP-A1 consisted of xylose, arabinose, glucose, mannose and galactose and ESP-A2 consisted of xylose, arabinose, rhamnose and galactose, in a molar ratio (%) of 3.2: 61.1: 11.1: 12.9: 11.6 and 20.6: 67.7: 5.0: 6.7, respectively. The molecular weights (Mw) of ESP-A1 and ESP-A2 were 5.83 × 104 Da and more than 200 × 104 Da, respectively. To the best of our knowledge, two neutral polysaccharides are now being reported for the first time in this study.

Functional characteristics of non-starch polysaccharides (NSP) obtained from native (n) and malted (m) finger millet (ragi, Eleusine coracana, indaf-15)

Non-starch polysaccharides (NSP) were isolated from a recently released, fast germinating hybrid variety of finger millet and its malt (ragi, Indaf-15), in different yields and their functional characteristics, such as viscosity, oxidative gelation capacity, foam stabilization, and their effects on various dough properties, were studied. Viscosities of hemicellulose-B (hemi-B) fractions, obtained from native and malted ragi flours, was found to be greater (N – 3.04 and M – 1.98 at 1% concentration) than all other NSP fractions. A linear increase in viscosity of this fraction was noticed with respect to its concentration; however it decreased as the temperature increased from 30 to 80 °C and it was found to be maximum at pH 6.0. Hemi-B was found to stabilize protein foams against thermal disruption and showed a high water absorption (4.4 ml at 1% concentration) compared to cold water-soluble polysaccharides (CWSP) (3.0 ml at 1% concentration). Minor variations in dough characteristics, such as dough development time (DDT), dough stability (DS), tolerance index (TI), maximum extensibility (Em) and maximum resistance (Rm) were noticed upon the addition of hemi-B polysaccharides obtained from native and malted ragi and also CWSP (N). However, a significant decrease in the above parameters was noticed upon the addition of CWSP (M). Similar patterns were also noticed in starch pasting characteristics, such as gelatinization temperature (GT), peak viscosity (PV), cold paste viscosity (CPV), hot paste viscosity (HPV), breakdown viscosity (BV) upon the addition of CWSP (N) and hemi-B (N&M). However, a visible decrease in the above parameters was noticed upon the addition of CWSP (M). Oxidative gelation studies indicated minor variations in their flow properties before and after the addition of hydrogen peroxide and peroxidase. The results obtained from this present study indicated that ragi can be incorporated as a source of dietary fibre both in the native and malted forms, in the preparation of various health foods without altering the dough characteristics or the quality of the end-product.

Studies on the water soluble constituents of lichens. II. Antitumor polysaccharides of Lasallia, Usnea, and Cladonia species.

From six species of lichens, a variety of polysaccharide fractions were prepared and their antitumor effects were tested against the sarcoma 180 implanted in mice. These fractions were characterized by chemical and physicochemical methods. The active principle of Lasallia pensylvanica has been determined as GE-3 type glucan (a partially O-acetylated β-(1→6)-glucan). Another highly effective β-glucan, containing (1→3)-and (1→4)-linkages in the ratio 3 : 7, was isolated from Usnea rubescens, and it has been identified with lichenin. In addition, presence of so far unknown heteropolysaccharides with moderate antitumor effect has been suggested in the minor fraction of the lichen. Four Cladonia lichens, i.e.C. crispata, C. mitis, C. rangiferina subsp. grisea, and C. squamosa, were also examined, and it has been revealed that, in all of them, complex heteroglycans consisting chiefly of mannose, galactose, and glucose occur commonly as predominant, cold-water soluble polysaccharides, together with a small amount of α-glucan presumed to be PC-3 type (an α-glucan possessing (1→3)- and (1→4)-linkages alternately). Both the heteroglycan fraction and the glucan fraction of C. squamosa were shown to exert moderate antitumor effect.

Carbohydrate composition of tissues infected by the autoecious rusts, Puccinia pelargonii-zonalis and P. malvacearum

The following fractions—ethanol soluble sugars, cold water soluble polysaccharides, hot water soluble polysaccharides and insoluble polysaccharides in both rust-infected and uninfected leaves of Pelargonium inquinans (L.) Ait. infected with Puccinia pelargonii-zonalis Doidge, were analysed by means of gas-liquid chromatography. Ethanol soluble sugars of Althaea rosea Cav. infected by Puccinia malvacearum Mont. were also studied. The soluble carbohydrate distribution in the infected leaves indicates that the rust fungus stores certain acyclic polyols (arabitol and mannitol) and trehalose. Polysaccharides such as a cold water soluble glucan and a mannan are stored at the site of the rust pustules.

Polysaccharides of lichens and fungi. V. Antitumour active polysaccharides of lichens of Evernia, Acroscyphus and Alectoria spp.

The polysaccharides of some lichens were studied chemically in relation to their host-mediated antitumour activities against sarcoma 180 in mice. A water-insoluble polysaccharide fraction, EP-3, [α]17D +200°, from Evernia prunastri (L.) ACH. was shown to be an α-glucan containing α(1→3)(1→4) linkages in the ratio of 4 : 1. The cold water-soluble fraction, EP-4, was separated into EP-7, [α]17D +12°, and EP-6, [α]17D +164°. The former was shown to be an antitumour active glucan having β (1→3) (1→4) linkages in the ratio of 3 : 1, while the latter was revealed to be an α(1→3) (1→4) glucan (approx, ratio 3 : 2). The properties of these polysaccharides were discussed in comparing with the data of everniin reported by micovic, et al. A polysaccharide AS-3-IS, [α]14D+176°, isolated from Acroscyphyus sphaerophoroides LEV. giving an intensive blue iodine-colour reaction was shown to be a new type of α-glucan consisting of α(1→3), (1→4) and (1→6) linkages. A cold water-soluble polysaccharide, AS-3-S, was revealed to be a new glycopeptide. The polysacchrides, ASu-2 and ASa-2 isolated from Alectoria sulcata (LEV.) NYL. and A. sarmentosa (ACH.) ACH. were proved to be lichenin, and ASu-3 and ASa-3 were shown to be isolichenin. The host-mediated antitumour activities of the polysaccharide fractions isolated from the above lichen species were tested.

Studies on the Carbohydrates of Black Tea

The carbohydrates in black tea were divided into fourteen fractions. Free sugars and the component sugars of each fraction have been investigated mainly by the method of paper chromatography and the following results were obtained (see Table 2).Fructose, glucose and arabinose were detected as free monosaccharides, and sucrose, maltose, raffino-e and stachyose as free oligosaccharides.Inositol.and two kinds of unknown substances as free sugaralchols were also detected.Rhamnose, glucose, galactose, fructose, xylose and galacturonic acid were detected as the components of glycosides.Arabinose, galactose, galacturonic acid, rhamnose and glucose were detected as the components of hot 50% ethanol-soluble polysaccharides.Galactose, galacturonic acid, arabinose, rhamnose, ribose (?), de_oxyribose (?) and glucuronic acid were detected as the components of cold water-soluble polysaccharides, and besides this arabinose, galactose, galacturonic acid, desoxyribose, ribose, glucuronic acid, rhamnose and mannuronic acid as the components of hot water-soluble polysaccharides.Arabinose, galactose, galacturonic acid and small amounts of glucuronic acid and mannuronic acid were detected as the components of hot 0.5%, ammonium oxalate-soluble polysaccharides.Xylose, mannose, glucose, ribose, desoxy ribose, galactose and mannuronic acid as the components of hemicellulose, galactose, rhamnose, desoxyribose, xylose and ribose as the β-cellulose, xylose, galactose and rhamnose as the y-cellulose components were detected.a-Cellulose and a small amount of lignin were also detected.

花びらの炭水化物に関する研究

The carbohydrates in the staminate petal of Cucurbita moschata DUCH, pumpkin, were investigated (see Table 2). The following results were obtained. Fructose, glucose, sucrose, raffinose (?), stachyose (?) and one unknown spot were detected as free sugars. Glucose, fructose, galactose, arabinose, rhamnose and xylose were detected as the components of glycosides. Galactose, arabinose, galacturonic acid, glucose, xylose, rhamnose, fructose, ribose (?) and desoxyribose were those of the hot 50% ethanol-soluble polysaccharides. Glucose, galactose, arabinose and galacturonic acid were the components of cold water-soluble polysaccharides, and in addition, glucose, arabinose, galacturonic, acid, ribose and desoxyribose of hot water-soluble polysaccharides. Galactose, arabinose, galacturonic acid and glucose were found as the component sugars of hot 0.5% ammonium oxalate aq.-soluble polysaccharides (protopectin fraction). Glucose, mannose, arabinose, rhamnose, fructose, ribose and desoxyribose were as those of hemicellulose. Cellulose and a small amount of lignin were also found.

花びらの炭水化物に関する研究

The carbohydrates in the petals of Camellia japonica L., camellia, were studied (see Table 2). The following results were obtained. Glucose, fructose, sucrose, maltose, and two unknown spots were detected as the free sugars. Galactose, glucose, rhamnose, arabinose, ribose, fructose, mannose, galacturonic acid (?) and mannuronic acid (?) were detected as the components of glycosides. Glucose, galactose, arabinose, fructose and galacturonic acid were found to be the component sugars of hot ethanol-soluble polysaccharides. Arabinose, galacturonic acid, glucose and galactose as the components of the cold water-soluble polysaccharides, and more, arabinose, ribose, glucose and galacturonic acid were found as those of hot water-soluble polysaccharides, respectively. Arabinose galactose and galacturonic acid were those of the protopectin fraction (hot 0.5% ammonium oxalate aq.-soluble polysaccharides). Xylose, rhamnose, arabinose, ribose, mannose, galactose, galacturonic acid, glucuronic acid, glucose and desoxyribose were the components of hemicellulose. α-Cellulose and a small amount of lignin were found as usual.