Perspectives in the Application of High, Medium, and Low Molecular Weight Oat ß-d-Glucans in Dietary Nutrition and Food Technology-A Short Overview.

For centuries human civilization has cultivated oats, and now they are consumed in various forms of food, from instant breakfasts to beverages. They are a nutrient-rich food containing linear mixed-linkage (1 → 3) (1 → 4)-ß-d-glucans, which are relatively well soluble in water and responsible for various biological effects: the regulation of the blood cholesterol level, as well as being anti-inflammatory, prebiotic, antioxidant, and tumor-preventing. Numerous studies, especially in the last two decades, highlight the differences in the biological properties of the oat ß-d-glucan fractions of low, medium, and high molecular weight. These fractions differ in their features due to variations in bioavailability related to the rheological properties of these polysaccharides, and their association with food matrices, purity, and mode of preparation or modification. There is strong evidence that, under different conditions, the molecular weight may determine the potency of oat-extracted ß-d-glucans. In this review, we intend to give a concise overview of the properties and studies of the biological activities of oat ß-d-glucan preparations depending on their molecular weight and how they represent a prospective ingredient of functional food with the potential to prevent or modulate various pathological conditions.

Mid-Infrared Spectroscopic Study of Cultivating Medicinal Fungi Ganoderma: Composition, Development, and Strain Variability of Basidiocarps.

Attenuated total reflection Fourier-transform infrared (ATR-FTIR) spectroscopy was proposed for rapid, versatile, and non-invasive screening of Ganoderma basidiocarps to assess their potential for specific applications. Fifteen species and strains of this fungus were selected for analysis, and fine sections at different parts of young and mature basidiocarps were obtained. The spectra of fungal samples showed significant differences interpreted in terms of biochemical composition using characteristic bands of proteins, polysaccharides, lipids, and triterpenoids. Obviously, for the transverse sections in trama, especially in the basal part, the most intense bands at 950-1200 cm-1 corresponded to polysaccharide vibrations, while for the superficial sections, the bands of carbonyl and aliphatic groups of triterpenoids at 1310-1470, 1550-1740, and 2850-2980 cm-1 predominated. The pilei, especially hymenium tubes, apparently contained more proteins than the bases and stipes, as evidenced by the intense bands of amide vibrations at 1648 and 1545-1550 cm-1. The specificity of the Ganoderma basidiocarp is a densely pigmented surface layer rich in triterpenoids, as proved by ATR-FTIR spectroscopy. The spectral differences corresponding to the specificity of the triterpenoid composition may indicate the prospects of individual strains and species of this genus for cultivation and further use in food, cosmetics, or medicine.

Fractionation of the water insoluble part of the heterotrophic mutant green microalga Parachlorella kessleri HY1 (Chlorellaceae) biomass: Identification and structure of polysaccharides.

The water-insoluble part of Parachlorella kessleri HY1 biomass was subjected to the extraction of cell-wall polysaccharides using polar aprotic solvents (DMSO, LiCl/DMSO) and aqueous alkaline solutions (0.1, 1 and 4 mol·l-1 of NaOH). Proteins predominated in all the crude extracts and in the insoluble residues were partially removed by treatment with proteolytic enzymes (pepsin and pronase), and in some cases with the HCl/H2O2 reagent, yielding purified polysaccharide-enriched fractions. These treatments led to the solubilisation of some products in water. The composition and structure of isolated polysaccharides were characterised based on monosaccharide composition, glycosidic linkage and spectroscopic analyses. The DMSO extract contained mainly proteins, and polysaccharides were not detected. The water-soluble parts isolated from the LiCl/DMSO extract contained α-l-rhamnan, α-d-glucan and ß-d-glucogalactan; the water-insoluble part contained (1 â†’ 4)-ß-d-xylan, first isolated from the biomass of green microalgae. The alkali extracts contained polysaccharides of similar structure, and also water-insoluble (1 â†’ 4)-ß-d-mannan. The insoluble part after all extractions contained α-chitin as the main polysaccharide, which was confirmed by spectroscopic methods. All these polysaccharides can play a certain role in the cell wall structure of this microalga.

Polysaccharides from Basidiocarps of the Polypore Fungus Ganoderma resinaceum: Isolation and Structure.

In this study, we focused on the isolation and structural characterization of polysaccharides from a basidiocarp of polypore fungus Ganoderma resinaceum. Polysaccharide fractions were obtained by successive extractions with cold water at room temperature (20 °C), hot water under reflux (100 °C), and a solution of 1 mol L-1 sodium hydroxide. The purity of all fractions was controlled mainly by Fourier transform infrared (FTIR) spectroscopy, and their composition and structure were characterized by organic elemental analysis; neutral sugar and methylation analyses by gas chromatography equipped with flame ionization detector (GC/FID) and mass spectrometry detector (GC/MS), respectively; and by correlation nuclear magnetic resonance (NMR) spectroscopy. The aqueous extracts contained two main polysaccharides identified as a branched O-2-ß-d-mannosyl-(1→6)-α-d-galactan and a highly branched (1→3)(1→4)(1→6)-ß-d-glucan. Mannogalactan predominated in the cold water extract, and ß-d-glucan was the main product of the hot water extract. The hot water soluble fraction was further separated by preparative anion exchange chromatography into three sub-fractions; two of them were identified as branched ß-d-glucans with a structure similar to the corresponding polysaccharide of the original fraction. The alkaline extract contained a linear (1→3)-α-d-glucan and a weakly branched (1→3)-ß-d-glucan having terminal ß-d-glucosyl residues attached to O-6 of the backbone. The insoluble part after all extractions was identified as a polysaccharide complex containing chitin and ß-d-glucans.

Screening of the Chemical Composition and Identification of Hyaluronic Acid in Food Supplements by Fractionation and Fourier-Transform Infrared Spectroscopy.

Hyaluronic acid, together with collagen, vitamins or plant extracts, is a part of many cosmetic and food preparations. For example, this polysaccharide is used in formulation of many food supplements due to its protective effects on human health. In this work, the screening of the chemical composition of three chosen dietary supplements (powder, tablets and capsules) containing hyaluronic acid was carried out using Fourier-transform infrared spectroscopy. Because of the low amount of analyte in all these samples, it was isolated or concentrated prior to the analysis using a suitable sequential fractionation protocol. Individual isolation procedures were established for each sample based on their declared composition. Firstly, the major components such as collagen or vitamins were removed to obtain polysaccharide fractions by the enzymatic treatment and/or washing out with the appropriate solvents. In some cases, the water insoluble part was removed from the rest dissolved in water. Then, hyaluronic acid was precipitated with copper(II) cations and thus separated from the other polysaccharides. Finally, the analyte was identified in the enriched fractions by the characteristic vibrational bands. The amount of hyaluronic acid in the purified fractions was determined in three ways: gravimetrically, spectrophotometrically, and using isotachophoresis. The combination of the appropriate preparative and analytical steps led to the successful evaluation of chemical composition, finding and quantification of hyaluronic acid in all the studied samples.

Immunoactive polysaccharides produced by heterotrophic mutant of green microalga Parachlorella kessleri HY1 (Chlorellaceae).

Hot water extract from biomass of heterotrophic mutant green alga Parachlorella kessleri HY1 (Chlorellaceae) was deproteinised, and three polysaccharidic fractions were obtained by preparative chromatography. The low-molecular fraction (1.5 × 104g mol-1) was defined mainly as branched O-2-ß-xylo-(1→3)-ß-galactofuranan where xylose is partially methylated at O-4. Two high-molecular fractions (3.05 × 105 and 9.84 × 104g mol-1) were complex polysaccharides containing α-l-rhamnan and xylogalactofuranan parts in different ratios. The polysaccharides were well soluble in hot water and, upon cooling, tended to self-segregate. Immunomodulatory activities of the obtained fractions were preliminary tested using ELISA, FACS and ImmunoSpot kits. The polysaccharides increased the TNF-α production in melanoma bearing mice with much higher intensity than in healthy mice. This was in agreement with the FACS results on T and B cells indicating their possibly secondary activation by innate immunity cells.

Expanding the phenotype spectrum associated with pathogenic variants in the COL2A1 and COL11A1 genes.

We report the clinical findings of 26 individuals from 16 unrelated families carrying variants in the COL2A1 or COL11A1 genes. Using Sanger and next-generation sequencing, 11 different COL2A1 variants (seven novel), were identified in 13 families (19 affected individuals), all diagnosed with Stickler syndrome (STL) type 1. In nine families, the COL2A1 disease-causing variant arose de novo. Phenotypically, we observed myopia (95%) and retinal detachment (47%), joint hyperflexibility (92%), midface retrusion (84%), cleft palate (53%), and various degrees of hearing impairment (50%). One patient had a splenic artery aneurysm. One affected individual carrying pathogenic variant in COL2A1 showed no ocular signs including no evidence of membranous vitreous anomaly. In three families (seven affected individuals), three novel COL11A1 variants were found. The propositus with a de novo variant showed an ultrarare Marshall/STL overlap. In the second family, the only common clinical sign was postlingual progressive sensorineural hearing impairment (DFNA37). Affected individuals from the third family had typical STL2 signs. The spectrum of disease phenotypes associated with COL2A1 or COL11A1 variants continues to expand and includes typical STL and various bone dysplasias, but also nonsyndromic hearing impairment, isolated myopia with or without retinal detachment, and STL phenotype without clinically detectable ocular pathology.

Polysaccharides from Basidiocarps of Cultivating Mushroom Pleurotus ostreatus: Isolation and Structural Characterization.

Oyster mushrooms are an interesting source of biologically active glucans and other polysaccharides. This work is devoted to the isolation and structural characterization of polysaccharides from basidiocarps of the cultivated oyster mushroom, Pleurotus ostreatus. Five polysaccharidic fractions were obtained by subsequent extraction with cold water, hot water and two subsequent extractions with 1 m sodium hydroxide. Branched partially methoxylated mannogalactan and slightly branched (1→6)-ß-d-glucan predominated in cold- and hot-water-soluble fractions, respectively. Alternatively, these polysaccharides were obtained by only hot water extraction and subsequent two-stage chromatographic separation. The alkali-soluble parts originating from the first alkali extraction were then fractionated by dissolution in dimethyl sulfoxide (DMSO). The polysaccharide insoluble in DMSO was identified as linear (1→3)-α-d-glucan, while branched (1→3)(1→6)-ß-d-glucans were found to be soluble in DMSO. The second alkaline extract contained the mentioned branched ß-d-glucan together with some proteins. Finally, the alkali insoluble part was a cell wall complex of chitin and ß-d-glucans.

SD-OCT imaging as a valuable tool to support molecular genetic diagnostics of Usher syndrome type 1.

A girl with profound congenital deafness and balance problems was found at 3.5 years of age to be a carrier of two novel compound heterozygous mutations in MYO7A that were predicted to be disease-causing. She also carried one known pathogenic mutation and one rare variant in USH2A. Fundus examination performed at 4.75 years revealed one small peripheral pigment deposit in the right eye, indicating probable retinal degeneration. Spectral domain optical coherence tomography (SD-OCT) showed a loss of photoreceptors throughout the macular area, except for the foveolar region, clearly confirming a diagnosis of Usher syndrome type 1. This case demonstrates that SD-OCT may be easily used in young children to confirm retinal disease, quantify the extent of retinal damage, and monitor disease progression.

Distinction of fungal polysaccharides by N/C ratio and mid infrared spectroscopy.

A set of fungal polysaccharide samples was characterised by elemental analysis and FTIR spectroscopy and compared with reference chitins, chitosans and ß-D-glucans. The nitrogen to carbon (N/C) values and FTIR spectra were used to compare the samples based on their composition. It was found that the N/C ratio correlates well with deacetylation degree (DD) of chitosans and chitin/glucan ratio R(chit) of fungal chitin ­ ß-D-glucan complexes with the exception of some samples having significant nitrogen and/or carbon admixtures. FTIR spectroscopy was indicative for the N-acetylation of chitins (chitosans) as well as for the chitin (chitosan) contribution to fungal polysaccharide preparations. Multivariate analyses of the FTIR data (HCA, PCA) discriminated samples and reference materials into several clusters depending on their similarity. Chitosan lactates, chitosan ­ ß-D-glucans and chitin ­ ß-D-glucans of high and low amounts of chitin were successfully discriminated from the reference polysaccharides and from each other. The proposed procedures based on the N/C ratio and multivariate analyses of FTIR spectra may be used in screening fungal polysaccharide preparations.

Preparation of amidated derivatives of carboxymethylcellulose.

Carboxymethylcellulose (CMC) was selected as substrate for amidation based on previous results described for monocarboxy cellulose (MCC) with the aim to prepare highly substituted products. In comparison with MCC containing uronic carboxyl groups at C-6 position, O-carboxymethyl groups in CMC should be more accessible for reagents because they are more distant from the polysaccharide chain. Two-step way of amidation was based on the esterification of CMC carboxyls by reaction with methanol and further amino-de-alkoxylation (aminolysis) of the obtained methyl ester with amidation reagents (n-alkylamines, hydrazine and hydroxylamine). Purity and substitution degree of the products were monitored by the vibration spectroscopic methods (FTIR and Raman) and organic elemental analysis. Analytical methods confirmed the preparation of highly or moderately substituted N-alkylamides, hydrazide and hydroxamic acid of CMC.

Cholesterol and fat lowering with hydrophobic polysaccharide derivatives.

Hydrophobic derivatives of highly methylated citrus pectin, chitosan and cellulose were prepared and tested as potential cholesterol lowering agents. Elemental analysis and spectroscopic methods confirmed high substitution degree for all of them. Substitution with long alkyl/acyl groups led to significant changes in physical and thermal properties of modified polysaccharides. Sorption of cholate and cholesterol by these polysaccharide-based sorbents was estimated in comparison with the synthetic drug cholestyramine. It was found that modified polysaccharides have high affinity to cholesterol. By contrast, cholestyramine was effective only in cholate sorption.

Cell wall components of Leptosphaeria maculans enhance resistance of Brassica napus.

Preparations with elicitation activity were obtained from the mycelium of Leptosphaeria maculans , a fungal pathogen of oilseed rape (Brassica napus). Crude delipidated and deproteinized extract from fungal cell walls induced expression of pathogenesis related gene 1 (PR1), hydrogen peroxide accumulation, and enhanced resistance of B. napus plants toward infection by L. maculans. Elicitation activity significantly decreased after treatment of a crude extract with α- or ß-glucanase. Monosaccharide composition analysis of a crude extract purified by ion-exchange chromatography revealed glucose (∼58 mol %), mannose (∼22 mol %), and galactose (∼18 mol %) as the major sugars. FT-IR and NMR spectra confirmed the presence of both carbohydrate and polypeptide components in the purified product. Correlation NMR experiments defined trisaccharide bound to O-3 of serine residue α-D-Glcp-(1→2)-ß-D-Galf-(1→6)-α-D-Manp-(1→3)-L-Ser. Terminal α-D-Glcp and (1→6)-ß-D-glucan were also detected. The obtained results strongly support the conclusion that these carbohydrates induce defense response in B. napus plants.

Determination of phytic acid and inositolphosphates in barley.

Phytic acid (PA) and lower inositolphosphates (InsP(n) ) is the main storage form of phosphorus in grains or seeds. The content of PA and InsP(n) in different varieties of barley was analyzed by capillary isotachophoresis and online-coupled capillary isotachophoresis with CZE. The electrolytes (in demineralized water) for the isotachophoretic analysis consisted of 10 mM HCl, 14 mM glycylglycine, and 0.1% 2-hydroxyethylcellulose (leading) and 10 mM citric acid (terminating). The optimized electrolytes for the online coupling isotachophoresis with zone electrophoresis analysis were mixtures of 5 mM HCl, 7 mM glycylglycine, and 0.1% 2-hydroxyethylcellulose (leading), 20 mM citric acid, 10 mM glycylglycine, and 0.1% 2-hydroxyethylcellulose (background) and 10 mM citric acid (terminating). PA and all studied InsP(n) were separated within 25 min and detected by a conductivity detector. Simple sample preparation (acidic extraction), sufficient sensitivity, speed of analysis, and low running cost are important attributes of the electrophoretic methods. The method was used for the determination of PA and InsP(n) in barley varieties within an ongoing research project.

Infrared spectroscopy and outer product analysis for quantification of fat, nitrogen, and moisture of cocoa powder.

The combination of the near infrared (NIR) and Fourier-transform infrared (FTIR) absorbance spectra (1100-2500nm and 4000-600cm(-1)) of 100 cocoa powder samples was used to build calibration models for the determination of the content of fat, nitrogen, and moisture. The samples that comprised the dataset had an average composition of 13.51% of fat, 3.77% nitrogen, and 3.98% moisture. The fat content ranged from 2.42 to 22.00%, the nitrogen from 0.88 to 4.48%, and moisture from 1.60 to 7.80%. For NIR, the relative root mean square error of cross-validation (RMSECV) was 7.0% (R(2)=0.96) for fat, 1.7% (R(2)=0.98) for nitrogen, and 5.2% (R(2)=0.94) for moisture. For FTIR, the relative RMSECV was 10.4% (R(2)=0.94) for fat and 3.9% (R(2)=0.95) for nitrogen. However, for moisture, it was not possible to build a calibration model with suitable predictability. The combination of the NIR and FTIR domains (data fusion) by outer product analysis PLS1 allowed to predict these parameters and to characterise frequencies in one domain based on the information of the other domain. This work allows to conclude that the second derivative of NIR is the recommended procedure to quantify fat, nitrogen, and moisture content in cocoa powders by infrared spectroscopy.

The complexation of metal cations by D-galacturonic acid: a spectroscopic study.

Solid complexes of D-galacturonic acid (GalA) with cobalt(II), copper(II), nickel(II) and oxovanadium(IV) (1-4) were prepared and characterised. The metal-to-ligand molar ratio was 1:2 for complexes 1-3 and 1:1 for complex 4. The alpha- and beta-anomers of GalA were detected in all the complexes in solid state and in solutions. An addition of small amounts of the paramagnetic complexes to the D2O solution of pure ligand led to NMR line broadening of some 1H and 13C nuclei. This broadening was sensitive to the anomeric state of GalA in the case of complexes 1 and 4. NMR and vibrational spectroscopic data indicate the formation of carboxylate complexes of all the cations, while noncarboxylic oxygens are also involved into the metal bonding in some cases. VCD spectra of complexes 1-4 in D2O and Me2SO-d6 solutions confirm that GalA carboxylic group may participate in the formation of optically active species around the metal cation. Possible ways of GalA coordination by metal cations of this study were proposed and discussed.