Rhamnose Content Research Articles

Pectins from Aloe Vera: Extraction and production of gels for regenerative medicine

ABSTRACTExtraction, purification, and gel preparation of Aloe Vera pectin and the evaluation of the biocompatibility of the pectin gels were studied, considering as end use as implantable materials for regenerative medicine. A. Vera was chosen as source of pectin, as this pectin was described to possess high molecular weight and a low degree of esterification. As the properties of pectins are strictly dependent upon the extraction methods in combination with the natural source, the extraction method was modified in order to optimize the yield of the final product, its purity, the duration of the process and the selection of non‐toxic chemical reagents. Changing the experimental conditions resulted in four different extraction processes and products with different physical and chemical characteristics. The optimal extraction resulted to be the process: with enzimatic deactivation by microwave and the use of sodium citrate as chelating agent the molecular weight of the pectin extracted was estimated to be 118 kDa and the 2.93% esterification degree. Cytocompatibility of pectin gels, prepared by ionotropic gelation, showing an improved cell adhesion if compared to commercial pectin. The results suggest that the extracted A. Vera pectins possess interesting properties to be exploited for the production of mechanically stable gels by ionotropic gelation and high rhamnose content matrices for application in regenerative medicine. © 2013 Wiley Periodicals, Inc. J. Appl. Polym. Sci. 2014, 131, 39760.

Modification of cell wall polysaccharide during ripening of Chinese bayberry fruit

Experiments were conducted to investigate the modification of cell wall polysaccharide in Chinese bayberry during ripening. Cellular wall materials were isolated from Chinese bayberry fruit at four ripening stages. Three pectin fractions and two hemicellulose fractions were obtained and their compositional changes were analyzed. The process of ripening and softening of the fruit corresponded to an increase in the water soluble pectin (WSP) and 1mol/L KOH-soluble hemicellulose (HC1) fractions, and a decrease in Na2CO3-soluble pectin (SSP) and 4mol/L KOH-soluble hemicellulose (HC2) fractions which suggested the solubilization and conversion of cellular wall components. However, the content of CDTA-soluble pectin (CSP) changed slightly. Furthermore, arabinose, as the major branch monosaccharide, decreased in three pectin fractions during the whole ripening period, whereas the relative content of rhamnose, the backbone monosaccaride, tend to decrease in CSP and SSP pectin fractions, but increased gradually in WSP pectin during ripening. Analysis of (Ara+Gal)/Rha ratio showed that the degradation of WSP pectin mainly occurred in branch chains, conversely, the degradation of backbone played a more important role in degradation of CSP and SSP. In HC1, the breakdown of the branch chain of matrix xyloglucan, arabinoxylan and arabinogalactan are predominant while more backbones of xyloglucan, arabinoxylan and arabinogalactan were depolymerized in HC2. Moreover, it was found that the molecular weight of WSP decreased gradually throughout ripening while the depolymerization of ionic CSP and covalent SSP mainly occurred at late and early ripening stages, respectively. Loosely bound (HC1) and tightly bound (HC2) hemicellulose fractions exhibited distinct molecular mass downshift throughout ripening stages, especially at late ripening stage. Overall, fruit softening of Chinese bayberry during ripening involved in differential modification of pectin and hemicellulose.

Insights into the Ropy Phenotype of the Exopolysaccharide-Producing Strain Bifidobacterium animalis subsp. lactis A1dOxR

The proteome of the ropy strain Bifidobacterium animalis subsp. lactis A1dOxR, compared to that of its nonropy isogenic strain, showed an overproduction of a protein involved in rhamnose biosynthesis. Results were confirmed by gene expression analysis, and this fact agreed with the high rhamnose content of the ropy exopolysaccharide.

Tamarind Seed Xyloglucans Promote Proliferation and Migration of Human Skin Cells through Internalization via Stimulation of Proproliferative Signal Transduction Pathways.

Xyloglucans (XGs) of Tamarindus indica L. Fabaceae are used as drug vehicles or as ingredients of cosmetics. Two xyloglucans were extracted from T. indica seed with cold water (TSw) and copper complex precipitation (TSc). Both were analyzed in regard to composition and influence on cell viability, proliferation, cell cycle progression, migration, MAPK phosphorylation, and gene expression of human skin keratinocytes (NHEK and HaCaT) and fibroblasts (NHDF) in vitro. TSw and TSc differed in molecular weight, rhamnose content, and ratios of xylose, arabinose, galactose, and glucose. Both XGs improved keratinocytes and fibroblast proliferation, promoted the cell cycle, and stimulated migration and intracellular enzyme activity of NHDF after endosomal uptake. Only TSw significantly enhanced HaCaT migration and extracellular enzyme activity of NHDF and HaCaT. TSw and TSc predominantly enhanced the phosphorylation of molecules that referred to Erk signaling in NHEK. In NHDF parts of the integrin signaling and SAPK/JNK pathway were affected. Independent of cell type TSw marginally regulated the expression of genes, which referred to membrane proteins, cytoskeleton, cytokine signaling, and ECM as well as to processes of metabolism and transcription. Results show that T. indica xyloglucans promote skin regeneration by a direct influence on cell proliferation and migration.

Improved Growth ofLactobacillus bulgaricusandStreptococcus thermophilusas well as Increased Antioxidant Activity by Biotransforming Litchi Pericarp Polysaccharide withAspergillus awamori

This study was conducted to increase the bioactivity of litchi pericarp polysaccharides (LPPs) biotransformed by Aspergillus awamori. Compared to the non-A. awamori-fermented LPP, the growth effects of A. awamori-fermented LPP on Lactobacillus bulgaricus and Streptococcus thermophilus were four and two times higher after 3 days of fermentation, respectively. Increased 1,1-diphenyl-2-picrylhydrazyl radical scavenging activity and DNA protection activity of litchi pericarp polysaccharides were also achieved after A. awamori fermentation. Moreover, the relative content of glucose and arabinose in LPP after fermentation decreased from 58.82% to 22.60% and from 18.82% to 10.09%, respectively, with a concomitant increase in the relative contents of galactose, rhamnose, xylose, and mannose. Furthermore, lower molecular weight polysaccharides were obtained after A. awamori fermentation. It can be concluded that A. awamori was effective in biotransforming LPP into a bioactive mixture with lower molecular weight polysaccharides and higher antioxidant activity and relative galactose content.

Role of polymer complexes in the formation of biofilms by corrosive bacteria on steel surfaces

The composition of exopolymer complexes (EPCs), synthesized by the monocultures Desulfovibrio sp. 10, Bacillus subtilis 36, and Pseudomonas aeruginosa 27 and by microbial associations involved in the corrosion of metal surfaces has been studied. An analysis of the monosaccharide composition of carbohydrate components, as well as the fatty acid composition of the lipid part of EPCs, was carried out by gas-liquid chromatography (GLC). It was found that bacteria in biofilms synthesized polymers; this process was dominated by glucose, while the growth of bacteria in a suspension was marked by a high rhamnose content. Hexouronic acids and hexosamine have been revealed as a part of B. subtilis 36 and P. aeruginosa 27 EPCs. Qualitative differences were revealed in the fatty acid composition ofexopolymers in biofilms and in a bacterial suspension. It was shown that the transition to a biofilm form of growth led to an increase in the unsaturation degree of fatty acids in the exopolymers of associative cultures. The results can be used to develop methods to control microbial corrosion of metal surfaces.

Adaptative biochemical pathways and regulatory networks in Klebsiella oxytoca BAS-10 producing a biotechnologically relevant exopolysaccharide during Fe(III)-citrate fermentation

BackgroundA bacterial strain previously isolated from pyrite mine drainage and named BAS-10 was tentatively identified as Klebsiella oxytoca. Unlikely other enterobacteria, BAS-10 is able to grow on Fe(III)-citrate as sole carbon and energy source, yielding acetic acid and CO2 coupled with Fe(III) reduction to Fe(II) and showing unusual physiological characteristics. In fact, under this growth condition, BAS-10 produces an exopolysaccharide (EPS) having a high rhamnose content and metal-binding properties, whose biotechnological applications were proven as very relevant.ResultsFurther phylogenetic analysis, based on 16S rDNA sequence, definitively confirmed that BAS-10 belongs to K. oxytoca species. In order to rationalize the biochemical peculiarities of this unusual enterobacteriun, combined 2D-Differential Gel Electrophoresis (2D-DIGE) analysis and mass spectrometry procedures were used to investigate its proteomic changes: i) under aerobic or anaerobic cultivation with Fe(III)-citrate as sole carbon source; ii) under anaerobic cultivations using Na(I)-citrate or Fe(III)-citrate as sole carbon source. Combining data from these differential studies peculiar levels of outer membrane proteins, key regulatory factors of carbon and nitrogen metabolism and enzymes involved in TCA cycle and sugar biosynthesis or required for citrate fermentation and stress response during anaerobic growth on Fe(III)-citrate were revealed. The protein differential regulation seems to ensure efficient cell growth coupled with EPS production by adapting metabolic and biochemical processes in order to face iron toxicity and to optimize energy production.ConclusionDifferential proteomics provided insights on the molecular mechanisms necessary for anaeorobic utilization of Fe(III)-citrate in a biotechnologically promising enterobacteriun, also revealing genes that can be targeted for the rational design of high-yielding EPS producer strains.

Influence of viscous Rhodella grisea (Rhodophyceae) proteoglycan on chemically induced cough reflex

An algal extracellular biopolymer (over 8.5×105Da) composed of carbohydrates (52%) and protein (∼13%) has been isolated from a red alga Rhodella grisea growing in natural conditions by concentration of water medium, alcohol precipitation, dialysis and freeze-drying. This mucilagineous biopolymer contained xylose and its 3-O- and 4-O-methyl derivatives (∼63%), galactose (∼12%), glucuronic acid (11–12%), glucose (∼5%), rhamnose (∼4%), fucose (∼3–4%) and low content of others accompaning sugars. When tested on the citric acid-induced cough and reactivity of airways smooth muscle in vivo in the test system guinea pigs, this biopolymer assigned a significant cough suppressing effect. The reactivity of airways smooth muscle was not affected indicating that expectoration effect was not suppressed by biopolymer application, which is important from the pharmacological point of view.

Enzymatic changes in pectic polysaccharides related to the beneficial effect of soaking on bean cooking time

Cooking time decreases when beans are soaked first. However, the molecular basis of this decrease remains unclear. To determine the mechanisms involved, changes in both pectic polysaccharides and cell wall enzymes were monitored during soaking. Two cultivars and one breeding line were studied. Soaking increased the activity of the cell wall enzymes rhamnogalacturonase, galactanase and polygalacturonase. Their activity in the cell wall was detected as changes in chemical composition of pectic polysaccharides. Rhamnose content decreased but galactose and uronic acid contents increased in the polysaccharides of soaked beans. A decrease in the average molecular weight of the pectin fraction was induced during soaking. The decrease in rhamnose and the polygalacturonase activity were associated (r = 0.933, P = 0.01, and r = 0.725, P = 0.01, respectively) with shorter cooking time after soaking. Pectic cell wall enzymes are responsible for the changes in rhamnogalacturonan I and polygalacturonan induced during soaking and constitute the biochemical factors that give bean cell walls new polysaccharide arrangements. Rhamnogalacturonan I is dispersed throughout the entire cell wall and interacts with cellulose and hemicellulose fibres, resulting in a higher rate of pectic polysaccharide thermosolubility and, therefore, a shorter cooking time.

Molecular characteristics of sulfated polysaccharides from Monostroma nitidum and their in vitro anticancer and immunomodulatory activities

We investigated water-soluble sulfated polysaccharides isolated from Monostroma nitidum using ion-exchange chromatography to determine their molecular characteristics and biological activities. The crude and fractionated polysaccharides (F 1, F 2, and F 3) consisted mostly of carbohydrates (58.3–91.9%), uronic acids (0–21.8%) and sulfates (1.8–17.7%) as well as varying amounts of proteins (1.6–9.4%). Their monosaccharide levels were significantly different including rhamnose (0–95.7%) and glucose (0–98.6%) content with small amounts of xylose (0.8–4.3%). These polysaccharides contained one or two subfractions with average molecular weights ( M w) ranging from 94.4 to 1387 × 10 3 g/mol. The in vitro inhibitory activity (≤75%) of the polysaccharides on a human cancer cell line (AGS) suggested that the polysaccharides had direct cytotoxic effects on the cancer cells. In addition, these hetero-polysaccharides (from the crude and F 1 and F 2 fractions) stimulated a macrophage cell line, Raw 264.7 cells, inducing considerable NO and PGE 2, production, which suggested that they could be strong immunomodulators.

Isolation and chemical characterization of algal polysaccharides from the green seaweed Ulva clathrata (Roth) C. Agardh

In order to obtain information on the content and composition of the water-soluble polysaccharides from Ulva clathrata, an extraction at 60°C, in different media, was performed: water, EDTA and HCl (F-I), each followed by a sequential extraction in NaOH 0.1 M (F-II). The extracts were recovered and analyzed for total carbohydrates, proteins, rhamnose, uronic acids and sulfate content. Differences were obtained in the yield and composition in both fractions of the different media (F-I and F-II). Higher yields resulted in the first fraction on all media. HCl extraction was the best in both fractions (14.83 ± 1.5% and 5.96 ± 1.1%, F-I and F-II, respectively). In all cases, F-I was more sulfated ranging from 27.87% to 35.8% and higher in rhamnose content, whereas F-II had higher protein and slightly higher uronic acid content. FTIR spectra showed that soluble polysaccharides from the green seaweed U. clathrata are sulfated polysaccharides, similar to ulvan obtained from other Ulva species and confirmed by the 1 H-NMR spectrum, where the characteristic signal for the deoxy sugar (rhamnose) is present.

α-galactosidase activity and carbohydrate mobilization in seeds of Dalbergia nigra (vell.) Allemão ex Benth. - fabaceae (Brazilian rosewood) during germination

Complex substances are converted by enzyme action into soluble molecules during the germination process, and these are in turn translocated to the growing plant embryo, serving as an energy source or physical structure. With the objective of quantifying α-galactosidase enzyme activity as well as mono-and oligosaccharide mobilization during germination, this study was conducted using Brazilian rosewood seeds. Seeds were kept in a germinator at 25ºC under continuous light for evaluation of radicle emergence (protrusion) over a period of 10 days. Mono-and oligosaccharide contents and also the specific activity of α-galactosidase enzyme were quantified on days zero, one, three and five. There was mobilization of glucose and mannose stored in cotyledons, and also of xylose in cotyledons and in the embryo in the first three days of germination. Rhamnose contents increased in cotyledons and in the embryonic axis. Raffinose was the initially used oligosaccharide, both in the embryonic axis and in cotyledons, while sucrose accumulated in both. Enzyme activity varied throughout, with greater specific activity on day one of imbibition, both in cotyledons and in the embryonic axis.

Loss of IrpT Function in Lactococcus lactis subsp. lactis N8 Results in Increased Nisin Resistance

The antibiotic nisin, produced by Lactococcus lactis subsp. lactis N8, offers an extensive commercial prospect as natural food preservatives. The nisin immunity of the L. lactis strains is regulated by a variety of mechanisms. In this study, we isolated a L. lactis L31 strain with increased nisin resistance from a mini-Mu transposon mutant pool of strain N8. The single Mu insertion in strain L31 was in the irpT gene with unknown function. By comparing the proteomic profiles of L. lactis L31 and its parental strain, we found that changes occurred in the synthesis of a protein involved in cell wall biosynthesis (RmlD). Strain L31 had 13.7% higher content of rhamnose in the cell wall than the N8 strain. Overexpression of RmlD involved in the synthesis of dTDP-L: -rhamnose in the nisin-sensitive MG1363 strain increased nisin resistance of the strain. The results indicate that these cellular proteins effected nisin resistance in L. lactis N8.

THE BIOLOGICAL PROPERTIES OF SOYBEAN NODULE BACTERIA WITH THE DIFFERENT SPEED OF GROWTH

The phenotypical and genotypic properties of soybean microsymbionts with the different speed of growth were studied. It was established that the strains with the intensive growth were specific to the host-plant, but had differed by their serological, chemotaxonomical and symbiotic properties. The investigated intensive-growing strains were combined into the one serogroup by their antigenic content. The similarity of fatty acids spectrum was observed for the strains with the intensive growth Bradyrhizobium sp. КВ1-1, slow-growing strains B. japonicum 634b and B. japonicum КС2-3. The quantitative and qualitative differences in monosaccharides contents of EPS of the studied strains were revealed. Presence of xylose (2,7 %) and increased contents of rhamnose (18,4 %) were shown to be different in EPS of strain Bradyrhizobium sp. КВ1- 1 and EPS of strains B. japonicum КС2-3 and B. japonicum 634b. Analysis of the sequences of the 16S rRNA genes allowed attributing the strain with the intensive growth to the Bradyrhizobium genus. Genes consistency analysis of 16S rRNA had allowed to refer the intensivegrowing strains to the Bradyrhizobium genus. It was shown that studied intensive-growing strains had formed the nonspecific symbiosis with cow pea.

Cell wall modification during development of mealy texture in the stony-hard peach “Odoroki” treated with propylene

The melting peach cultivar “Akatsuki” [ Prunus persica L. Batsh] develops melting texture during ripening, but the stony-hard peach cultivar “Odoroki”, softens and develops a mealy texture when treated with propylene. Cell wall polysaccharides were fractionated as 1,2-diaminocyclohexanetetraacetic acid (CDTA)-, Na 2CO 3-, NaOH-soluble fractions, or the cellulosic residues that remained after solvent extraction. Size-exclusion chromatography of the CDTA-soluble fraction revealed a limited depolymerization of polyuronides in mealy “Odoroki” fruit, accompanying a lower level of endo-polygalacturonase (endo-PG) and a higher level of exo-PG activities compared to melting “Akatsuki” fruit. “Akatsuki” fruit showed increases in the arabinose, galactose and rhamnose contents in the CDTA-soluble fraction. These neutral sugars decreased in the Na 2CO 3-soluble and cellulosic residue fractions during ripening. Similar changes were observed for neutral sugars other than arabinose during softening in mealy “Odoroki” fruit. Arabinose content did not increase in the CDTA-soluble fraction of “Odoroki” fruit; however, arabinose content decreased in the Na 2CO 3-soluble fraction and in the cellulosic residue as observed for “Akatsuki” fruit. A significant increase in α- l-arabinofuranosidase activity, about 10 times higher than for “Akatsuki” fruit, was observed in propylene-treated mealy “Odoroki” fruit. Differences in ripening-associated metabolic changes in the cell walls of these peach cultivars seem to be related to their differences in texture.

Comparative Study of the Physicochemical Properties of Rice Endosperm Components Expressing Sugary-2 Mutant in Different Genetic Backgrounds

The physicochemical properties of the endosperm components of sugary-2 mutant rice were examined, characterized and compared to that of the normal type. The protein and amylose contents as well as the palatability score were all higher in sugary-2. For fatty acids, palmitic, linoleic and linolenic acid were all present in significantly higher proportion in the sugary-2 mutant. Analysis of the minor sugars in the endosperm showed that inducing the sugary-2 gene did not increase the amount of all sugar types, higher amounts were only measured in rhamnose and fucose content. Regarding the characteristics of starch granules, the ratio of longer glucose chain length to the distribution of endosperm starches decreased with the insertion of the sugary-2 mutant gene. And it was also found that the gelatinization properties of sugary-2 mutant rice showed a general trend of lower gelatinization temperature while the gelatinization enthalphy was considerably lower.

Evaluation of fruit authenticity and determination of the fruit content of fruit products using FT-NIR spectroscopy of cell wall components

An analytical procedure using Fourier transform near infrared (FT-NIR) spectroscopy and chemometrics with multivariate techniques for the rapid determination of the fruit authenticity and for the quantification of the fruit content was developed, based on the cell wall constituents (alcohol-insoluble residue, AIR, and hemicellulose, HC). The contents of rhamnose, fucose, arabinose, xylose, mannose, galactose, and glucose in the hemicellulose fraction of apricots, peaches, and pumpkins determined by gas chromatography were used as references. Furthermore, spectral information was correlated with the fruit content and the gravimetric data obtained from sequential fractionation of the alcohol-insoluble residue. Samples of self-made and commercial apricot and peach fruit preparations, jams, and spreads were included in the investigations. Hemicellulose from 109 samples and AIR from 92 samples was recorded, and principal component regression was used to create calibration models relating chemical and gravimetrical reference values to spectral data. The calibration models provided a good predictability in comparison with the results obtained by reference methods. Good agreement was also obtained for the prediction of the neutral sugar composition of the HC and the fruit content from the AIR. FT-NIR spectroscopy allowed a rapid, accurate and non-destructive assignment of specified fruit from spectral data of the HC fraction and the AIR. Thus, FT-NIR could be applied for investigations on quality control complementing, or even replacing, gas chromatography as the most widespread method for the determination of neutral sugars. Furthermore, preliminary investigation on classification of fruit blends was performed. The probability of belonging to a specific fruit was significantly lowered or precluded for samples with fraudulent admixture like peach or pumpkin to apricot, or pumpkin to peach.

Effects of organic and inorganic amendments on soil organic matter properties

The aim of this work was to investigate the effects of long-term application of different organic fertilizers (sewage sludge, farmyard manure, compost) as compared to mineral fertilizer on the structure of the soil organic matter. Capillary electrophoresis was employed for the quantification of monosaccharides and phenolic compounds, whereas NMR and FT-IR were used for the overall characterization of the soils organic matter. Application of farmyard manure results in a higher content of organic matter derived from angiosperms, suggested by the higher levels of syringic and vanillic phenols. Spectroscopic studies show an increase of lignin and lignin-like products in the organic matter of the soil, which may be derived from the cereal straw supplied with farmyard manure. According to spectroscopic analysis, 13C CPMAS-NMR and FT-IR spectra, higher contents of methylene groups (– CH 2) from proteins and protein-like compounds, as well as higher levels of carbohydrates, were found in the soil supplied with compost. The monosaccharide (rhamnose, xylose, glucose, mannose, arabinose, fucose and galactose) content was not significantly influenced by the different organic fertilizers, suggesting that the type of amendment used does not affect any of these six studied monomers. Comparing the three organic amendments the most significant differences were observed after long-term application of farmyard manure, with an increase in lignin and lignin-like products in the soil organic matter, and compost, which appears to contribute to an increase of protein and protein-like, as well as carbohydrates content on soil organic matter. The knowledge of such changes can be essential to understand the sorption and bioavailability of pollutants, as well as establish/unestablish ways to recycling organic residues as organic fertilizers.

Overexpression of a cytosol-localized rhamnose biosynthesis protein encoded by Arabidopsis RHM1 gene increases rhamnose content in cell wall

l-Rhamnose (Rha) is an important constituent of pectic polysaccharides, a major component of the cell walls of Arabidopsis, which is synthesized by three enzymes encoded by AtRHM1, AtRHM2/AtMUM4, and AtRHM3. Despite the finding that RHM1 is involved in root hair formation in Arabidopsis, experimental evidence is still lacking for the in vivo enzymatic activity and subcellular compartmentation of AtRHM1 protein. AtRHM1 displays high similarity to the other members of RHM family in Arabidopsis and in other plant species such as rice and grape. Expression studies with AtRHM1 promoter-GUS fusion gene showed that AtRHM1 was expressed almost ubiquitously, with stronger expression in roots and cotyledons of young seedlings and inflorescences. GFP::AtRHM1 fusion protein was found to be localized in the cytosol of cotyledon cells and of petiole cells of cotyledon, indicating that AtRHM1 is a cytosol-localized protein. The overexpression of AtRHM1 gene in Arabidopsis resulted in an increase of rhamnose content as much as 40% in the leaf cell wall compared to the wild type as well as an alteration in the contents of galactose and glucose. Fourier-transform infrared analyses revealed that surplus rhamnose upon AtRHM1 overexpression contributes to the construction of rhamnogalacturonan.

Heparinoid-active two sulfated polysaccharides isolated from marine green algae Monostroma nitidum

Two sulfated polysaccharides WF1 and WF3 were isolated from marine green algae Monostroma nitidum, and their structural characteristics were determined. Anticoagulant activities of WF1 and WF3 were evaluated by assays of the activated partial thromboplastin time (APTT), thrombin time (TT), prothrombin time (PT), antithrombin and anticoagulation factor Xa activities. The results showed that WF1 and WF3 had similar high contents of rhamnose, whereas their sulfate contents, sulfation positions, molecular sizes and linkage patterns of rhamnose residues were different. The bioassay results demonstrated that WF1 and WF3 had high anticoagulant activities, and were potent thrombin inhibitors mediated by heparin cofactor II, especially WF3. They also hastened thrombin and coagulation factor Xa inhibition by potentiating antithrombin III, but at a lower effectiveness. The differences of anticoagulant activities between WF1 and WF3 were directly due to their structural features discrepancy.