Hot Water-soluble Fractions Research Articles

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-β-d-mannosyl-(1→6)-α-d-galactan and a highly branched (1→3)(1→4)(1→6)-β-d-glucan. Mannogalactan predominated in the cold water extract, and β-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 β-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)-β-d-glucan having terminal β-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 β-d-glucans.

Response of Different Crop Cultivars to Micronutrient Fertilization and Relationship to Rhizosphere Soil Properties

ABSTRACT Five different cultivars each of wheat, pea, and canola were grown with and without copper (Cu), zinc (Zn), and boron (B) fertilization, respectively, in a greenhouse study to identify any crop cultivar and rhizosphere soil responses to micronutrient fertilization. The prairie agricultural surface soils (0–10 cm) used in the study were suspected to be micronutrient deficient according to soil extractable levels. Only durum wheat (CDC Fortitude) produced greater biomass from Cu fertilization compared to the other wheat cultivars. Cultivar dependent responses of pea and canola biomass yield to Zn and B fertilization were not significantly (P > .05) different. Boron concentration in whole plant canola tissue increased following fertilizer addition. Micronutrient fertilization increased concentrations and supply rates of available Cu, Zn, and B in post-harvest rhizosphere soil. Fertilizer Cu and Zn were distributed in labile soil solution-carbonate-exchangeable and oxyhydroxide fraction, whereas B was primarily found in the hot water-soluble fraction, specifically adsorbed and oxyhydroxide fractions. The composition of low molecular weight organic acids (formic, propionic, malic, and fumaric acid) in the rhizosphere soil varied among crops. Additionally, citric and tartaric acid were present in the pea and canola rhizosphere soils, respectively. The rhizosphere soil of durum wheat not fertilized with Cu contain higher levels of succinic acid compared to the other wheat cultivars. For pea and canola, there was no significant (P > .05) effect of cultivar on the amounts or forms of organic acids in the rhizosphere soils.

Antitumor and Immunostimulating Activities of Elfvingia applanata Hot Water Extract on Sarcoma 180 Tumor-bearing ICR Mice

Elfvingia applanata, a medicinal mushroom belonging to Basidiomycota, has been used in the effort to cure cancers of the esophagus and stomach, and is also known to have inhibitory effects on hepatitis B virus infection. The hot water soluble fraction (as Fr. HW) was extracted from fruiting bodies of the mushroom. In vitro cytotoxicity tests showed that hot water extract was not cytotoxic against cancer cell lines such as Sarcoma 180, HT-29, HepG2, and TR at concentrations of 10~2,000 μg/mL. Intraperitoneal injection with Fr. HW resulted in a life prolongation effect of 45.2% in mice previously inoculated with Sarcoma 180. Treatment of Fr. HW resulted in a 2.53-fold increase in the numbers of murine spleen cells at a concentration of 50 μg/mL, compared with control. Incubation of murine spleen cells with Fr. HW at a concentration of 500 μg/mL resulted in improved immune-potwntiating activity of B lymphocytes through an 8.3-folds increase in alkaline phosphatase activity, compared with control. Fr. HW generated 12.5 μM of nitric oxide (NO) when cultured with RAW 264.7, a mouse macrophage cell line, at the concentration of 50 μg/mL, while lipopolysaccharide, a positive control, produced 15.2 μM of NO. Therefore, the results suggested that antitumor activities of Fr. HW from E. applanata might, in part, be due to host mediated immunostimulating activity.

Composting in small laboratory pilots: Performance and reproducibility

Small-scale reactors (<10 l) have been employed in composting research, but few attempts have assessed the performance of composting considering the transformations of organic matter. Moreover, composting at small scales is often performed by imposing a fixed temperature, thus creating artificial conditions, and the reproducibility of composting has rarely been reported. The objectives of this study are to design an innovative small-scale composting device safeguarding self-heating to drive the composting process and to assess the performance and reproducibility of composting in small-scale pilots. The experimental setup included six 4-l reactors used for composting a mixture of sewage sludge and green wastes. The performance of the process was assessed by monitoring the temperature, O 2 consumption and CO 2 emissions, and characterising the biochemical evolution of organic matter. A good reproducibility was found for the six replicates with coefficients of variation for all parameters generally lower than 19%. An intense self-heating ensured the existence of a spontaneous thermophilic phase in all reactors. The average loss of total organic matter (TOM) was 46% of the initial content. Compared to the initial mixture, the hot water soluble fraction decreased by 62%, the hemicellulose-like fraction by 68%, the cellulose-like fraction by 50% and the lignin-like fractions by 12% in the final compost. The TOM losses, compost stabilisation and evolution of the biochemical fractions were similar to observed in large reactors or on-site experiments, excluding the lignin degradation, which was less important than in full-scale systems. The reproducibility of the process and the quality of the final compost make it possible to propose the use of this experimental device for research requiring a mass reduction of the initial composted waste mixtures.

Structural differences between hot-water-soluble and hot-water-insoluble fractions of starch in waxy rice ( Oryza sativa L.)

Amylose readily dissolves in hot water, unlike amylopectin which is largely insoluble. However, size distributions of amylose isolated in such a manner often show the presence of hyper-branched material consistent with amylopectin. The difference in solubility suggests that the hot-water-soluble (HWS) hyper-branched material might be structurally distinct from typical amylopectin. In the present paper, the structural properties of the two solubility fractions of hyper-branched material are explored in a set of traditional waxy rice varieties. The objective was to elucidate the nature of the HWS component, e.g. to see if it could be phytoglycogen, another water-soluble polysaccharide. We show that solubility is controlled by thermodynamic effects, rather than slow dissolution (kinetic effects). The average size, degree of branching and the debranched chain-length distributions indicate that the HWS fraction is structurally different from phytoglycogen. The debranched chain-length distributions of short chains and the degrees of branching in the HWS material are similar to those of the hot-water-insoluble (HWI) fractions but the chain-length distributions indicate that the HWI fractions carry longer chains than those in the HWS fractions. Light-scattering measurements show that the average size of whole molecules in the HWI component is significantly greater than in the HWS component. It is postulated that the structural differences limit solubility of the molecules in the HWI fraction, possibly due to co-crystallisation with adjacent molecules at more points than is possible for the shorter chains in HWS molecules.

Effects of carbohydrate substrate on the vegetative mycelial growth of an ectomycorrhizal mushroom, Tricholoma matsutake, isolated from Quercus

We studied the characteristics of the utilization of carbohydrate substrates and the production of those hydrolyzing enzymes of the Tricholoma matsutake J-1 strain isolated from hardwood (Quercus sp.). In the culture medium, 5% glucose inhibited mycelial growth. The growth inhibition rate was remarkable in the Z-1 strain from softwood (Pinus densiflora) compared with that of the J-1 strain from hardwood. α-Amylase production varied with starches from different origins in contrast to mycelial growth. The range of the effect of 0.5%–15% soluble starch on vegetative mycelial growth was also investigated. The optimal concentration for mycelial growth was 15% for the J-1 strain but 10% for the Z-1 strain. Mycelial growth of the J-1 strain was strongly inhibited in PMML medium containing Sunpeal- CP prepared from sulfite pulp softwood waste, but that of the Z-1 strain was not inhibited by Sunpeal-CP. Moreover, mycelial growth of the J-1 strain from Quercus sp. dramatically decreased with the addition of CNF-HWSF (hot watersoluble fractions from corn fiber) to the PMML and PDL medium. However, inhibition by CNF-HWSF was not shown in the Z-1 strain from P. densiflora.

Effects of carbohydrate substrate on the vegetative mycelial growth of an ectomycorrhizal mushroom, &lt;i&gt;Tricholoma matsutake&lt;/i&gt;, isolated from &lt;i&gt;Quercus&lt;/i&gt;

We studied the characteristics of the utilization of carbohydrate substrates and the production of those hydrolyzing enzymes of the Tricholoma matsutake J-1 strain isolated from hardwood (Quercus sp.). In the culture medium, 5% glucose inhibited mycelial growth. The growth inhibition rate was remarkable in the Z-1 strain from softwood (Pinus densiflora) compared with that of the J-1 strain from hardwood. α-Amylase production varied with starches from different origins in contrast to mycelial growth. The range of the effect of 0.5%–15% soluble starch on vegetative mycelial growth was also investigated. The optimal concentration for mycelial growth was 15% for the J-1 strain but 10% for the Z-1 strain. Mycelial growth of the J-1 strain was strongly inhibited in PMML medium containing Sunpeal- CP prepared from sulfite pulp softwood waste, but that of the Z-1 strain was not inhibited by Sunpeal-CP. Moreover, mycelial growth of the J-1 strain from Quercus sp. dramatically decreased with the addition of CNF-HWSF (hot watersoluble fractions from corn fiber) to the PMML and PDL medium. However, inhibition by CNF-HWSF was not shown in the Z-1 strain from P. densiflora.

Effect of the addition of fatty acids on rice starch properties

The effect of the addition of the two fatty acids (stearic and linoleic acid) on starch pasting, thermal properties and the leaching of amylose during the gelatinization process are presented. The addition of stearic acid (C18:0) at 1.0% level (w/w, on the basis of starch) significantly changed starch pasting characteristics in peak viscosity (P < 0.05), breakdown (P < 0.001) and time to peak viscosity (P < 0.001) as measured by RVA. In contrast, the addition of linoleic acid (C18:2) showed less impact on the three RVA parameters. Thermal studies with DSC suggested that the addition of both fatty acids did not significantly influence the starch gelatinization behaviour as reflected in peak width and enthalpy but there was a decrease in peak temperature. However, the addition of the saturated fatty acid produced a significant decrease in the retrogradation endotherm (P < 0.01) compared to that of native starch. It is suggested that the saturated fatty acid complexes with amylose and is preferentially introduced into the starch granule. The double bond in the linoleic acid apparently hinders complexation. After complexation, hydrophobicity was further enhanced with the content of amylose in the hot-water soluble fraction being significantly reduced (P < 0.01) as determined by HPLC.

Improved Methods for the Structural Analysis of the Amylose-Rich Fraction from Rice Flour

Cooking and sensory properties of rice are largely determined by the amylose content and structure. For relationships between functional and structural properties, a more accurate method to determine the structure of amylose is required. Here we calibrate size exclusion chromatography (SEC) columns, using Mark-Houwink parameters for linear starch and pullulan standards, to obtain the true molecular weight distribution of linear starch. When the molecular weight distribution is reported relative to pullulan, rather than the actual molecular weight which is readily obtained from universal calibration, it is seen that the molecular weights of longer amylose chains are greatly underestimated. We validate the SEC method to enable the measurement of the hydrodynamic volume distribution of the starch by examining reproducibility and recovery. Analysis of the starch in the sample pre- and post-SEC shows that 20% of the carbohydrate is not recovered. Comparison of the weight-average degree of polymerization, X(w), of (undebranched) starch of pre- and post-SEC is made using iodine binding as well as Berry plots of data from multi-angle laser light scattering (MALLS). These both show that current SEC techniques for starch analysis lead to significant loss of high molecular weight material. Indeed, for the systems studied here, the values for X(w) after SEC are about three times lower than those before SEC. Iodine-starch complexes of pre- and post-SEC samples reveals that the SEC techniques give reliable data for the amylose fraction but not for amylopectin. We address reports in the literature suggesting that the conventional isoamylase method for debranching starch would lead to incomplete debranching and thus incorrect molecular weight distributions. However, it is shown using (1)H NMR that isoamylase can completely debranch the amylose (to within the detection limit of 0.5%), and by SEC that successive incubation with isoamylase, alpha-amylase, and beta-amylase can degrade the amylose-rich fraction completely to maltose. We develop a method to obtain a hot water soluble fraction (HWSF), rich in undamaged amylose molecules, directly from rice flour, avoiding the structural degradation of previous techniques. With appropriate sample handling, the formation of associations between starch chains is minimized. With the combination of calibrated and validated SEC methods, and an improved extraction of amylose from rice, the X(w) for both HWSF and debranched HWSF are found to be much larger than has previously been reported.

The stimulation of extracellular carbohydrases of edible mushrooms by the hot water-soluble fraction from corn fiber

Corn fiber (CNF) is an abundant by-product of the wet corn milling process in the production of cornstarch. We have shown that the hot water-soluble fraction (HWSF) from CNF has a promoting effect on the mycelial growth of various edible mushrooms, including mycorrhizal fungi. To reveal the promoting mechanisms, the effect of CNFHWSF on the stimulation of extracellular enzymes was examined. The production of extracellular carbohydrases such as amylase, CMCase, and xylanase was markedly enhanced by the addition of low molecular weight fractions (less than MW 500) prepared from CNF-HWSF. The enzymatic stimulation and enhancement of mycelial growth appeared during 3–15 days after inoculation. Furthermore, a fraction of less than MW 500 was separated by gel filtrate chromatography into five fractions (A–E), and the effect of each fraction was investigated. Promoting effects were shown from C and D fractions; mycelial growth and enzyme production of Lentinula edodes were indicated although fraction D has no sugars and amino acids in CNF-HWSF. From these results, the promoting effect of CNF-HWSF seems to be a two-step reaction. The first step could be achieved by rich nutrients such as free amino acids and monosaccharides from CNF-HWSF. The second step (during 3–15 days) is considered to be that the marked promoting effect was caused by the stimulation of extracellular enzymes.

Promotive effect of the hot water-soluble fraction from corn fiber on vegetative mycelial growth in edible mushrooms

Corn fiber (CNF) is an abundant by-product of the wet corn milling process used to produce corn starch. In light of the need to recycle organic wastes, the effects of adding a hot water-soluble fraction (HWSF) from CNF to a medium on the vegetative mycelial growth of nine edible mushrooms such as Lentinula edodes and Pholiota nameko were investigated. The results showed that the mycelial growth of these fungi was markedly increased (1.4–9.5 times that of the control) by adding 5%–20% CNF-HWSF to the medium. These promotive effects were also apparent on mycorrhizal mushrooms, such as Tricholoma matsutake (3.3-fold) and Lyophyllium shimeji (3.7-fold). The promotive effects on mycelial growth were shown in the low-molecular-weight fractions (molecular weight <500 daltons) prepared from CNF-HWSF. The promotive actions were more effective on slow-growing mushrooms (L. edodes and P. nameko) than on rapidly growing mushrooms (Pleurotus ostreatus and Flammulina velutipes). The results obtained in this experiment suggest that CNF-HWSF can be used as a promotive substance for cultivating edible mushrooms.

Polysaccharides from Agaricus blazei Stimulate Lymphocyte T-Cell Subsets in Mice.

Subset analysis of splenic lymphocytes using flow cytometry showed that the percentages of Thy1.2-(pan T-cells), L3T4-(CD4, helper T-cells), and Lyt2-(CD8, cytotoxic T-cells) positive cell populations were significantly increased in mice orally administered a hot water-soluble fraction from Agaricus blazei as compared with mice treated only with saline. 13C-NMR data indicates that the main component in the active polysaccharide is the complex of alpha-1,6- and alpha-1,4-glucan, which had already been shown to have anti-tumor activity against Sarcoma 180. It seems that the polysaccharide from Agaricus blazei may be an effective prophylactic, protecting humans against cancer by stimulating lymphocytes such as cytotoxic T-cells.

Carbohydrate composition in relation to structural stability, compactibility and plasticity of two soils in a long-term experiment

The effects of tillage on soil organic carbon content, carbohydrate content, monosaccharide composition, aggregate stability, compactibility and plasticity were investigated in a field experiment on a gleysol and on a cambisol under winter barley in South-East Scotland. Two long-term treatments (direct drilling and conventional mouldboard ploughing for 22 years) were compared with short-term direct drilling and broadcast sowing plus rotavation for 5 years. Carbohydrate released sequentially to cold water, hot water, 1.0 M HCl and 0.5 M NaOH was determined after hydrolysis as reducing sugar equivalent to glucose in both fresh and air-dried samples. All other measurements were made on dry soils only. About 3% of the soluble carbohydrate was extracted by cold water, 10% by hot water, 12% by HCl and 75% by NaOH from both the dry and fresh soils. The total reducing sugars of the fractions were proportional to the total organic carbon determined by dichromate oxidation or C analysis. Organic carbon and carbohydrates were concentrated near the surface of the direct drilled soil, but were more uniformly distributed with depth in the ploughed soil. The surface soil under direct drilling was more stable, less compactible and had greater plasticity limits than under ploughing. However, particle size distributions were unaffected by tillage so that differences in soil properties were attributed to differences in the quantity and quality of organic matter. Differences in compactibility, structural stability and plasticity limits between depths and tillage treatments correlated with total carbon and with total carbohydrates. The hot water extractable carbohydrate fraction correlated best with aggregate stability and the NaOH fraction correlated best with compactibility and plastic limit. Both fractions were greatest in the long-term direct drilled soil. The hot water fraction had a galactose plus mannose over arabinose plus xylose ratio of 1.0–1.6 in comparison to 0.4–0.7 in the NaOH fraction indicating that the microbial contribution within the hot water-soluble fraction was the greater. The hot-water fraction was likely to contain more exocellular microbial polysaccharides involved in the stabilizing of soil aggregates. The hot-water and NaOH carbohydrate fractions may be good indicators of soil organic matter quality relevant to the preservation of good soil physical conditions.

Differences in Immunomodulating Effects Between Wild and Cultured Panax ginseng

The different effects between wild and cultured Panax ginseng on immunological activity were investigated. The extracts of hot water soluble fraction from wild Panax ginseng showed the mitogenic activity to lymphocytes but that from cultured Panax ginseng did not. The mitogenic activity of wild Panax ginseng (100 μg/well) was almost equal to Concanavalin A (0.1 μg/well) which was well-known as one of T cell mitogens. The percentages of Thy1.2-(pan T cells), L3T4-(helper T cells) and Lyt2-(cytotoxic T cells) positive cell population were significantly increased in the mice orally administered hot water soluble fraction from wild Panax ginseng as compared to control by 31.2, 17.9 and 30.1 percent, respectively.

Chemical change in organic matter decomposing in and on a forest Rendzina under beech (Fagus sylvatica L.)

SUMMARYThe chemical characteristics of the organic matter of five horizons from a Terra fusca Rendzina under beech were compared by fractionation. Two litter layers, sampled in autumn, were compared: fresh leaf litter and the remains of the previous years’ litter. Three mineral horizons, A, B and C were also sampled.While decomposition in the litter layer led only to slight redistribution of the fractions, much greater changes occurred when the organic matter became incorporated into the mineral soil. The proportion of C and N in amino sugars rose markedly down the profile. The percentage of more readily soluble substances, such as the hot water‐soluble fraction or the unidentified, hydrolysable component, in the mineral soil was twice the amount found in the leaf litter. The percentage of non‐hydrolysable compounds in the mineral horizons was nearly half that found in the leaf litter. The C/N ratio fell from 44 in fresh leaf litter to 10 in the C horizon. The amino acid C/amino sugar C ratio fell from 45 in fresh leaf litter to 3.0 in the B horizon. The glucosamine/galactosamine ratio fell from infinite in galactosamine‐free fresh leaf litter to 1.2 in residual litter and increased again to 2.5 in the C horizon. The hexose/pentose ratios were virtually the same in all horizons. The glucose/ mannose ratio showed a steady fall down the profile, from 7.7 to 1.6. The glucuronic/ galacturonic acid ratio increased from 0.1 and 0.2 in the two litter layers to 4.7 in the C horizon.

A comparison between the hot and cold water soluble fractions of two locust bean gum samples

Locust bean gum extracted from two carob flours from eastern and western Mediterranean sources were fractionated on the basis of their solubility in water. Weight-average molecular weights determined by sedimentation equilibrium were about 300 000 for both the hot water and cold water soluble fractions, whereas a commercial sample of guar gum had a molecular weight of 700 000. Their values were lower than would be predicted from Mark-Houwink relationships where molecular weights were originally determined by light scattering. The hot water soluble fraction from the eastern Mediterranean flour showed unexpected rheological behaviour. It had an extremely high Huggins' constant and a different relationship between the coil overlap parameter and the zero shear rate viscosity compared with previously reported results for galactomannans. Both effects may be explained by the anomalously low intrinsic viscosity of this fraction when determined by a Huggins' extrapolation. The use of the Kraemer extrapolation gave significantly higher intrinsic viscosities for this particular sample. Gels formed from the two hot water soluble fractions with κ-carrageenan had similar rheological properties.

A simplified method for determining collagen in fish muscle.

The muscle of carp was fractionated into water-soluble, alkali-soluble (0.1N NaOH), acid-soluble (0.5M acetic acid), hot water-soluble (extraction with water at 120°C for 1h), and residual fractions. The analyses of amino acid and SDS-polyacrylamide gel electrophoresis showed that collagen in the muscle was quantitatively recovered only in the acid-soluble and hot water-soluble fractions and non-collagenous proteins were absent in these fractions. On the basis of the colori-metric determination of protein in these fractions, we devised a simplified method for determinating the soluble and insoluble collagens in the muscle. The soluble and insoluble collagens in the muscles of the rainbow trout Salmo gairdnerii, Japanese mackerel Scomber japonicus, carp Cyprinus carpio, and eel Anguilla japonica were deter-mined by the proposed method. The total collagen contents in the muscles of rainbow trout, Japanese mackerel, carp, and eel were found to be 0.47, 0.50, 0.60, and 1.99% of the wet muscle, respectively.

HEMICELLULOSES OF WHEAT STRAW

Various hemicellulose fractions were extracted from wheat straw holocellulose (extractive and pectin free) by successive treatments with cold and hot water, 0.5%, 1.0%, and 2.0% potassium hydroxide and were recovered by precipitation with alcohol. Approximately 25% of the holocellulose material was removed, one half being in the hot water soluble fraction. The original holocellulose, the extracted residue, and the recovered fractions were analyzed for pentosan, uronic acid anhydride, acetyl, methoxyl, and ash content. In general, the more soluble fractions had a higher uronic acid and methoxyl content; the less soluble had a higher pentosan content and a more negative rotation [Formula: see text]. Intrinsic viscosity measurements indicated that all fractions had a degree of polymerization of 25–30. Hydrolysis of the main fraction yielded D-xylose, L-arabinose, D-glucose; in addition D-galactose was found in the water soluble fractions. Quantitative determinations of the sugars in the hydrolyzates showed that D-xylose predominated, with L-arabinose, D-glucose, and D-galactose (when present) in progressively smaller amounts. On hydrolysis all fractions yielded an acid-resistant uronic acid complex that contained D-xylose and a uronic acid tentatively identified as monomethoxyl galacturonic acid.