Purification and identification of polysaccharide derived from Chlorella pyrenoidosa

The conditions for extracting and purifying polysaccharides from Chlorella pyrenoidosa, including intensity and duration of ultrasound, the temperature and incubation time, and ethanol concentration, were investigated through an orthogonal design of L₁₆(4⁵) in this work. High performance liquid chromatography (HPLC) and gas chromatography (GC) were used to characterize the compounds in C. pyrenoidosa. The highest yield of 44.8 g kg⁻¹ was achieved at 400 W of ultrasound for 800 s and then followed by incubation in water bath at 100 °C for 4 h in 80% ethanol. Two polysaccharide fractions (S1 and S2) were separated from the extracts of C. pyrenoidosa using Sepharose 4B column chromatography. The average molecular weights (M_w) of S1 and S2 were 81,877 Da and 1749 Da, respectively. Gas chromatographic (GC) traces of the hydrolyzed polysaccharides showed that most of the majority of monosaccharide in both fractions was mannose (78.0% and 76.5% of relative mass from S1 and S2, respectively) with low levels of glucose (13.2% and 8.4% of relative mass from S1 and S2, respectively). The Fourier-transform infrared spectra (FT-IR) of S1 and S2 revealed typical characteristics of polysaccharides. Both samples had the characteristics of hydroxyl groups, weak C–H band and α-pyranoses; however, only S2 had a carboxyl group.     

Chromatographic determination of polysaccharides in Lycium barbarum Linnaeus

Polysaccharides in Lycium barbarum Linnaeus have been shown to be effective in preventing cancer. The objectives of this study were to develop an appropriate method for molecular weight determination of polysaccharides in L. barbarum. The most suitable analytical condition was: a volume-ratio of L. barbarum sample to deionized water at 1:10, followed by shaking in a 100 °C water bath for 30 min, concentrating to 50 mL and adding 250 mL of 95% ethanol for precipitation at −20 °C for 8 h, hydrolysing protein with 2.5 U/mL of proteinase at pH 8 and 60 °C for 4 h and separating polysaccharide into five fractions by high-performance size exclusion chromatography (HPSEC) with the molecular weight of two major fractions being 79,250 and 24,468 Da. Analysis of monosaccharides by gas chromatography (GC) indicated the presence of rhamnose, arabinose, xylose, mannose, glucose and galactose, with the molar ratio at 0.3:2.7:0.3:0.2:2.7:0.9, respectively.     

Structural characterisation of a water-soluble polysaccharide with high branches from the leaves of Taxus chinensis var. mairei

A large number of polysaccharides present in the leaves of Taxus chinensis var. mairei were successively extracted with hot water after preliminary treatments. The ion-exchange and gel-permeation chromatography was used to isolate and purify the major polysaccharides to afford a complex water-soluble polysaccharide, named T1 with a molecular mass of 3.44 × 10⁶ Da determined by HPGPC. T1 consisted of 2,4-di-O-methyl-mannose, rhamnose, arabinose, xylose, galactose, mannose, galacturonic acid and glucuronic acid in a molar ratio of 2:5:24:9:3:46:1:10, and the acetyl content was estimated to be 2.1%. On the basis of methylation analysis, periodate and chromium trioxide oxidations, Smith degradation, graded acid hydrolysis, and NMR and IR spectroscopy, T1 possessed a 1,3,6-linked β-d-Manp main chain, substituted by mannose, arabinose, galactose, xylose, rhamnose, 2,4-di-O-methyl-mannose, galacturonic acid, and glucuronic acid residues. These contained non-reducing end-units of mannose, arabinose, xylose, 2,4-di-O-methyl-mannose, and glucuronic acid. This is the first report on isolation of an acid heteropolysaccharide from Taxus species.     

Purification,preliminary structural characterization and in vitro antioxidant activity of polysaccharides from Acanthus ilicifolius

Crude Acanthus ilicifolius (A. ilicifolius) polysaccharides (CAIP) were obtained by hot water extraction and deproteinated. Two major polysaccharide fractions, the neutral A. ilicifolius polysaccharide (NAIP) and the acid A. ilicifolius polysaccharide B (AAIP-B), were isolated from CAIP by chromatography using DEAE-Sepharose Fast Flow and Sepharose CL-6B. The molecular weights (M_w) of NAIP and AAIP-B, determined by high performance gel-filtration chromatography (HPGFC), were 11,775 and 23,161 Da, respectively. AAIP-B contained 51.23% uronic acid, characteristic of a pectin-type hetero-polysaccharide. Analysis of the neutral monosaccharide composition indicated that NAIP contained high proportions of arabinose, galactose and glucose. However, AAIP-B was mainly composed of rhamnose, arabinose and galactose. Their structure properties were confirmed by FT-IR.     

Three sulphated polysaccharides isolated from the mucilage of mud snail, Bullacta exarata philippi: Characterization and antitumour activity

Three sulphated polysaccharides, coded as BEMPA, BEMPB₁, BEMPB₂, were extracted from the mucilage of mud snail of Bullacta exarata and purified by DEAE-cellulose ion-exchange and size-exclusion chromatography. Structural analysis of purified polysaccharides by chemical and biochemical methods revealed BEMPA was a high (1→3,4)-linked mannose-containing polysaccharide with molecular weight of 22,977 Da. BEMPB₁, with molecular weight of 64,117 Da, was a high (1→3)-linked arabinose-containing polysaccharide. BEMPB₂ was mainly composed of (1→3,4)-linked mannose with molecular weight of 47,507 Da. The comparison between sulphated polysaccharides and their desulphated products showed that sulphate substitutions of BEMPB₁ were deduced to be at the C-3 of (1→4)-linked mannose, while sulphate substitutions of BEMPA and BEMPB₂ were at C-4 of (1→3)-linked mannose. Furthermore, BEMPA exhibited highest inhibitory effects on growth of B-16 melanoma cells, and IC₅₀ were 31.1 μg/mL.     

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

A new heteropolysaccharide with a molecular weight of 1.94 × 10⁴ Da, HEPF1, was isolated from the fruiting bodies of Hericium erinaceus. It is composed of fucose, galactose and glucose in the ratio of 1:4:1, as well as a minor proportion of 3-O-methyl rhamnose. Sugar analyses, methylation analysis, together with ¹H and ¹³C NMR spectroscopy established that HEPF1 has a (1 → 6)-linked α-d-galactopyranosyl backbone with branches that are composed of fucose attached to O-2; it also contains 6-O-substituted-β-d-oligoglucosyl units and a minor terminal 3-O-methyl rhamnose residue.     

自溶对双孢菇多糖体外抗氧化和抗肿瘤活性的影响

本研究旨在探究自溶对双孢菇多糖（Agaricus bisporus polysaccharides,ABP）的分子质量以及体外抗氧化和抗肿瘤活性的影响。通过对在不同温度和时间下发生自溶的双孢菇进行多糖提取,分离纯化具有强抗氧化和抗肿瘤活性的ABP组分,测定分子质量并分析单糖组成。结果表明,与不自溶组相比,ABP由于自溶降解,出现了更多的中分子质量多糖。自溶作用增强了低分质子量多糖的体外抗氧化和抗肿瘤活性。具有最佳抗氧化活性和体外抗肿瘤活性的多糖分子质量为619.72 Da,主要的单糖组成为葡萄糖、半乳糖和鼠李糖。     

Structural characterization of water-soluble polysaccharides from Opuntia monacantha cladodes in relation to their anti-glycated activities

An aqueous extract of polysaccharides from Opuntia monacantha cladodes (POMC) was preliminarily purified by 5 kDa molecular weight cut-off ultrafiltration membrane to remove impurities with low molecular weight. Then the retentate was fractionated by ethanol solution and chromatographed on a DEAE Sepharose Fast Flow anion-exchange column to yield a major fraction (POMC IV) which was eluted by 0.5 M NaCl. POMC IV was subjected to further purification on a Sephadex G-50 gel filtration column. Two major fractions, POMC V and VI, were collected. By analyses using gel permeation chromatography (GPC), high-performance liquid chromatography (HPLC) and gas chromatography (GC), POMC V, which had a molecular weight of 28.7 kDa, was comprised mainly of rhamnose, arabinose and glucose in the molar ratio of 9.15:1.00:6.84, with 3.07% (w/w) of glucuronic acid, while POMC VI, which had a molecular weight of 10.8 kDa, was comprised mainly of rhamnose, mannose and glucose in the molar ratio of 8.72:1.00:6.19, with 4.68% (w/w) of glucuronic acid. Six distinct-absorbance peaks, at 1742, 1633 and 1417 cm⁻¹ in the infrared (IR) spectra of POMC V, and at 1729, 1596 and 1407 cm⁻¹ in the IR spectra of POMC VI, resulted from the presence of uronic acids. The peaks at 1043 and 890 cm⁻¹ were characteristic of rhamonse and β-d-glucose, respectively. From the profiles of ¹³C and ¹H nuclear magnetic-resonance (NMR) spectra, the main (1 → 2)-α-l-rhamnopyranose units were obviously characterized by six strong signals at 99.24 (C-1), 77.52 (C-2), 70.19 (C-3), 71.33 (C-4), 69.81 (C-5) and 17.45 ppm (C-6). The signal at 175.92 ppm was due to C-6 of β-d-glucuronic acid units. The ¹H spectrum signal at 1.20 ppm was assigned to the CH₃ of α-l-rhamnopyranose units. The evaluation of anti-glycation activity suggested that POMC had good potential for inhibiting the formation of advanced glycation endproducts. Time- and dose-dependent effects were also observed for all POMC samples.     

Effect of exopolysaccharide-producing strains of Streptococcus thermophilus on technological attributes of fat-free lassi

Sixty-four exopolysaccharide-producing thermophilic lactic acid bacteria (LAB) were isolated from traditionally made Indian fermented milk products. On the basis of morphological and biochemical tests, these isolates were identified as the species of Lactobacillus, Streptococcus, and Enterococcus genera. Initial screening for technological attributes revealed that Streptococcus thermophilus IG16 was a promising isolate, and produced both capsular and ropy polysaccharides at the concentration of 211 mg/L. Exopolysaccharide produced by IG16 was a heteropolysaccharide containing rhamnose and galactose in a ratio of 5.3:1 and had a molecular weight of 3.3 × 10⁴ Da. Use of IG16 as a starter culture controlled whey separation and improved viscosity, flavor, consistency, and color and appearance of lassi. Use of IG16 resulted in lassi having optimal acidity, less syneresis, high viscosity, and better scores for flavor, consistency, and color and appearance.     

Separation and quantification of component monosaccharides of the tea polysaccharides from Gynostemma pentaphyllum by HPLC with indirect UV detection

A reversed-phase high-performance liquid chromatographic (HPLC) method is described for the simultaneous determination of aldoses and uronic acids. The separation was carried out on a RP-C₁₈ column (4.6 mm i.d. × 250 mm, 5 μm, Venusil, USA) using precolumn derivatization with 1-phenyl-3-methyl-5-pyrazolone (PMP) and UV detection at 250 nm, and the 10 PMP derivatives of mannose, ribose, rhamnose, glucuronic acid, galacturonic acid, glucose, xylose, galactose, arabinose and fucose were baseline separated within 40 min. Furthermore, the described method was applied to the quantitative analysis of component monosaccharides in the water-soluble polysaccharides extracted from Gynostemma pentaphyllum Makino tea and the result showed that the tea polysaccharide was a typical heteropolysaccharide and consisted of mannose, ribose, rhamnose, glucuronic acid, galacturonic acid, glucose, xylose, galactose and arabinose in the molar contents of 16.3, 10.3, 47.1, 5.6, 24.0, 128.4, 25.0, 101.4 and 71.1 μM, respectively. Quantitative recoveries of the component monosaccharides in the tea polysaccharide were in the range of 94.6–108.0% and the RSD values were lower than 4.9%. The results demonstrated that the proposed HPLC method was precise and practical for the analysis of the G. pentaphyllum tea polysaccharide.     

Hypocholesterolaemic effect of Bifidobacterium animalis subsp. lactis (Bb12) and trypsin casein hydrolysate

Hypercholesterolaemia is one of the most important risk factors in the development of cardiovascular disease. In this study, we report the in vitro potential of Bifidobacterium animalis subsp. lactis (Bb12) cultures and bovine casein hydrolysates formed by trypsin and Bb12 culture to reduce cholesterols levels. Cholesterol levels in vitro were reduced by up to 48% after incubation with Bb12 and up to 87% after incubation with trypsin hydrolysates, whereas unhydrolysed bovine casein did not affect cholesterol levels. Individual peptide fractions, obtained from size-exclusion chromatography, from casein hydrolysates formed by trypsin after a 48 h hydrolysis, reduced cholesterol levels by 2.7–50%. The molecular masses of these fractions, containing hypocholesterolaemic peptides, were below 1200 Da, as determined by LC-MS.     

Purification and characterisation of a hypoglycemic peptide from Momordica Charantia L. Var. abbreviata Ser.

A water-soluble peptide MC2-1-5 from Momordica charantia L. Var. Abbreviata Ser., with hypoglycemic effect, was purified by ultrafiltration, gel filtration chromatography and reverse-phase high performance liquid chromatography (RP-HPLC). The infrared (IR) spectra showed characteristic absorption peaks and the molecular mass of MC2-1-5 was found to be 3405.5174 Da by matrix-assisted laser desorption ionization time of flight mass spectrometry (MALDI-TOF-MS). The sequence of its first 10 N-terminal amino acids was GHPYYSIKKS as determined by a protein sequencer. MC2-1-5 reduced the blood glucose level in alloxan-induced diabetic mice by 61.70% and 69.18% at 2 and 4 h, respectively, after oral administration at a dose of 2 mg/kg. The oral glucose tolerance test (OGTT) showed MC2-1-5 produced a reduction of 25.50%, 39.62% and 41.74% in blood glucose level after 1, 2 and 3 h, respectively, of oral administration compared with a diabetic control.     

Preparation,preliminary characterization and immunostimulatory activity of polysaccharide fractions from the peduncles of Hovenia dulcis

The peduncles of Hovenia dulcis, containing abundant nutrients and having a taste like a combination of raisin, clove, cinnamon and sugar, have been consumed as fruits and used as traditional herbal medicine for a long time in China. Up to date, little information is available about the polysaccharides from peduncles of H. dulcis (HDPS) and their potential bioactivity. In this study, three purified fractions were prepared by sequential purification of crude HDPS through ion-exchange chromatography and gel-filtration chromatography. The three fractions of HDPS-1, HDPS-2 and HDPS-3 were found to be homogeneous heteropolysaccharides mainly composed of rhamnose, arabinose, galactose and galacturonic acid with an average molecular weight of 235, 70 and 53 kDa, respectively. HDPS-3 was quite different from HDPS-1 and HDPS-2, as it contained much higher content of galacturonic acid (40.5%). In vitro immunostimulatory activity evaluation revealed that all the three fractions could significantly stimulate the proliferation of splenocytes and enhance phagocytosis, nitric oxide production and acid phosphatase activity of peritoneal macrophages, which suggested that HDPS had a potent immunostimulatory activity and could be explored as a potential natural immunomodulatory agent.     

Antioxidant properties of polysaccharides from Ganoderma tsugae

Ganoderma tsugae Murrill (Ganodermataceae) were available in the form of mature and baby Ling chih, mycelia and fermentation filtrate. From these four forms, hot water extracted and hot alkali extracted polysaccharides were prepared and their antioxidant properties were studied. Polysaccharides showed good antioxidant activity as evidenced by their particularly low EC₅₀ values (<0.1 mg/ml). At 20 mg/ml, both extracted polysaccharides from mycelia showed reducing powers of 0.41–0.52 whereas reducing powers of other polysaccharides were in the range of 0.87 to 1.14. At 20 mg/ml, scavenging abilities on 1,1-diphenyl-2-picrylhydrazyl radicals increased to 93.7–100%, except for that of the hot water extracted polysaccharide, from filtrate, being 74.9%. At 20 mg/ml, scavenging abilities of both extracted polysaccharides from mycelia on hydroxyl radicals were 13.9 and 24.4%, respectively whereas scavenging abilities of the other polysaccharides were in the range of 39.0–55.2%. At 10 mg/ml, the chelating abilities of polysaccharides from mature and baby Ling chih, mycelia and filtrate were 93.9–100%, 97.6–100%, 85.1–88.0% and 51.2%, respectively. Overall, both extracts of polysaccharides possessed good antioxidant properties except for scavenging ability on hydroxyl radicals and can be developed as a new dietary supplement and functional food.     

Structural analysis and CCK-releasing activity of a sulphated polysaccharide from abalone (Haliotis Discus Hannai Ino) viscera

A fraction of water-soluble sulphated polysaccharide conjugate, termed AHP-2, was obtained from abalone (Haliotis Discus Hannai Ino) viscera by protease-assisted aqueous extraction followed by precipitation with ethanol and purification with gel filtration chromatography. Analysis by high performance liquid chromatography (HPLC) coupled to a TSK-gel G4000PW_XL column and gel filtration chromatography on Sepharose CL-6B indicated AHP-2 is homogenous with an average molecular weight (MW) of about 11.0 kDa. The structure of AHP-2 was revealed by chemical methods, Fourier transform infrared (FTIR) spectroscopy, and nuclear magnetic resonance (NMR) spectroscopy. Results indicated AHP-2 is a heteroglycan consisting of glucose, fucose, xylose, rhamnose and galactose with molar ratio of 1.0:2.0:3.9:6.7:7.4. The backbone of AHP-2 consists of 1,3-linked rhamnose and 1,3,6-linked galactose, with glucose, fucose, xylose and galactose of different linkage types distributing in branched chains. Meanwhile, AHP-2 was found to increase cholecystokinin (CCK) release in CCK-secreting STC-1 cells.     

Anti- and pro-oxidant activity of Cichorium genus vegetables and effect of thermal treatment in biological systems

The antiradical activity of water soluble components in six vegetables belonging to the Cichorium genus, i.e., three cultivars of red intybus species var. silvestre (Treviso, Chioggia, Verona red chicories), a white intybus species var. foliosum (Belgian chicory), and two vegetables of the endivia species var. latifolium (escarole chicory) and var. crispum (“crispa” chicory), were studied using two biological systems consisting of: (1) microsome membrane rat hepatocyties in which oxidative damage was induced by CCl₄; (2) gram-positive bacterium, Staphylococcus aureus cultures, subjected to damage with cumene hydroperoxyde. The obtained results show that in both systems the red vegetables possess the strongest antioxidant properties and contain different antioxidant compounds whether at a low or high molecular weight, but only those of high molecular-weight (MW > 3500 Da) are able to act as antioxidants in all the used systems. The lower MW fraction (MW < 3500 Da) showed itself to be pro-oxidant in the microsome system. The effects of thermal treatments such as boiling, freezing and freeze-drying were also investigated.     

Tenderness enhancement of beef from Bos indicus and Bos taurus cattle following electrical stimulation

High voltage electrical stimulation (1130 V peak, 14.28 bidirectional half sinusoidal pulses/s) or low voltage stimulation (45 V peak, 36 alternating square wave pulses/s) was used on cattle: (1) low voltage stimulation applied for 10 or 40 s with fast and slow chilling or high voltage stimulation for 60 s with normal chilling, applied to 100% Bos taurus cattle, (2) low voltage stimulation (40 s) and high voltage stimulation (60 s) with normal chilling applied to mixed Bos indicus and B.taurus cattle, (3) high voltage stimulation (54 s) with normal chilling applied to B. taurus and B. indicus cattle of 0–100% B. indicus composition, and (4) high voltage stimulation (60 s) applied to 100% B. taurus and 100% B. indicus cattle. All stimulation parameters enhanced the tenderness of steaks from M. longissimus thoracis et lumborum (LTL) aged at 1 °C up to 28 days compared with non stimulated LTL. Short low voltage stimulation of 10 s was marginally more effective than no stimulation and longer durations of 40 s were very effective and high voltage stimulation was most effective.     

Structure of a galactoarabinoglucuronoxylan from tamarillo (Solanum betaceum), a tropical exotic fruit,and its biological activity

Tamarillo (Solanum betaceum) is a tropical exotic fruit whose polysaccharides were extracted from the ripe pulp. After various purification steps, homogeneous fractions (designated PTW, STK-1000R and PF) were analyzed by sugar composition, HPSEC, methylation and NMR spectroscopy analysis. The results showed that the fraction PTW consisted of a linear arabinan with (1 → 5)-linked α-l-arabinofuranosyl units. Fractions designated as STK-1000R and PF contained galactoarabinoglucuronoxylans, with (1 → 4)-linked β-d-Xylp residues in the backbone, carrying branches exclusively at O-2. The polysaccharide in STK-1000R is less branched than that in the PF fraction (∼20.0% and 36.5%, respectively), with side-chains formed by (1 → 5)-linked α-l-Araf residues and (1 → 4)-linked α-d-GlcpA residues and with non-reducing end units formed by α-l-Araf, β-Arap, β-d-Galp, α-d-GlcpA and 4-O-Me-α-d-GlcpA. Intraperitoneal administration of the STK-1000R fraction in mice significantly reduced the number of abdominal constrictions induced by 0.6% acetic acid and the inflammatory phase of nociception induced by 2.5% formalin, indicating that that fraction has an antinociceptive effect on inflammatory pain models.     

Emulsifying and foaming properties of Phaseolus vulgaris and coccineus proteins

Protein isolates from two Phaseolus cultivars, common bean (Phaseolus vulgaris L.) and scarlet runner bean (Phaseolus coccineus L.), were prepared by wet extraction methods (isoelectric precipitation – 4000 rpm, ultrafiltration, extraction with NaCl 2%, and isoelectric precipitation – 9900 rpm). The protein isolates were characterized by sodium dodecyl sulfate–polyacrylamide gel electrophoresis and then evaluated for their solubility. The emulsion stability of emulsions produced at pH 7.0 and 5.5 with 1% or 2% or 3% w/v protein isolate was evaluated by average droplet size diameter, viscosity and creaming measurements. Emulsions with 1% protein content were unstable through storage. Emulsions with 3% w/v protein isolate concentration, extracted by ultrafiltration at pH 5.5 from both cultivars, were flocculated; this was more pronounced for coccineus isolates. The foaming properties, for the respective foams, were investigated. Foams with 1% w/v protein showed little foaming ability Ultrafiltration isolates produced more foam, which was especially stable at pH 5.5.