Thermally induced conformation transition of triple-helical lentinan in NaCl aqueous solution.

Lentinan, a beta-(1-->3)-D-glucan, was isolated from Lentinus edodes by using an improved extraction and purification method to show good water solubility and high yield. The results from 13C NMR, size-exclusion chromatography combined with multiangle laser light scattering (SEC-MALLS), dynamic light scattering (DLS), and optical rotation revealed that lentinan existed in a triple-helical conformation in the aqueous solution at 25 degrees C, whereas the thermally induced conformation transition from triple helix to single flexible chains occurred at elevated temperatures. The dependences of the weight-average molecular weight (Mw), radius of gyration (z1/2), hydrodynamic radius (Rh), intrinsic viscosity ([eta]), and specific optical rotation of lentinan on temperature in 0.9% NaCl aqueous solution showed an abrupt drop at 130-145 degrees C. It was confirmed that the conformation transitions from triple strand to single chain and from extended chains to winding chains for lentinan were completed rapidly at 130-145 degrees C, as a result of the simultaneous destruction of the intra- and intermolecular hydrogen bonds in lentinan. The thermally induced conformational transition was irreversible. The results from atomic force microscopy (AFM) and DLS demonstrated the existence of intrachain entanglement for the triple-helical chains, leading to the wormlike linear, circular, and crossover species for lentinan having high Mw (1.71x10(6)) in aqueous solution at 25 degrees C.

Structural characterization and antiviral activity of lentinan from Lentinus edodes mycelia against infectious hematopoietic necrosis virus.

A novel lentinan (LNT-I) was extracted from Lentinus edodes mycelia, and purified by an anion-exchange DEAE cellulose column and Sephadex G-200 gel. The structural characterization of LNT-I was determined by gas chromatography-mass spectrometry, high performance gel permeation chromatography, Fourier transform infrared spectrometry and 1D-nuclear magnetic resonance spectroscopy. The results showed that LNT-I was a ß-(1 → 3)-glucan backbone with -(1 → 6)-glucosyl side-branching units terminated by mannosyl and galactosyl residues, and its molecular weight was 3.79 × 105 Da. LNT-I consisted of glucose, mannose and galactose with the molar ratio of 19.26:1.20:1.00. LNT-I represented the prominent antiviral activity to IHNV at MOI of 0.05 and 0.10, respectively. Direct inactivation and the antiviral ability in pre-addition, co-addition and post-addition to IHNV (MOI of 0.05) were 62.34%, 39.60%, 53.63% and 82.38%, respectively under 100 µg/mL of LNT-I. Antiviral mechanisms of LNT-I mainly involved in the direct inactivation and the inhibition of viral replication. Moreover, LNT-I significantly down-regulated the expression level of TNF-α, IL-2 and IL-11, and up-modulated the expression levels of IFN-1 and IFN-γ after challenging with IHNV. The results indicated that the inhibitory effects of LNT-I on IHNV infection were possibly attributed to its regulation of the innate immune responses and specific immunity.

Optimised isolation of polysaccharides from Lentinula edodes strain NCBI JX915793 using response surface methodology and their antibacterial activities.

Mycelial growth in a defined medium by submerged fermentation is a rapid and alternative method for obtaining fungal biomass of consistent quality. Biomass, exopolysaccharides (EPS) and intracellular polysaccharides (IPS) production were optimised by response surface methodology in Lentinula edodes strain LeS (NCBI JX915793). The optimised conditions were pH 5.0, temperature 26°C, incubation period of 25 days and agitation rate of 52 r/min for L. edodes strain LeS. Under the calculated optimal culture conditions, biomass production (5.88 mg mL(-1)), EPS production (0.40 mg mL(-1)) and IPS production (12.45 mg g(-1)) were in agreement with the predicted values for biomass (5.93 mg mL(-1)), EPS (0.55 mg mL(-1)) and IPS production (12.64 mg g(-1)). Crude lentinan exhibited highest antibacterial effects followed by alcoholic, crude and aqueous extracts. The results obtained may be useful for highly effective yield of biomass and bioactive metabolites.

Optimization of selenizing conditions for Seleno-Lentinan and its characteristics.

Lentinan was successfully modified with nitric acid-sodium selenite method based on L9(3(4)) orthogonal experiments. The optimum selenizing conditions were obtained according to selenium conversion rate as follows: Lentinan of 1.0g, pH of 4.5, temperature of 70°C and sodium selenite of 1.50g. The antioxidant activity assays in vitro (DPPH, reducing power, superoxide radicals and hydroxyl radicals) proved that Lentinan had stronger antioxidant activity after selenizing. The elevations of serum alanine aminotransferase and aspartate aminotransferase, as well as the abnormal hepatic architecture, verified that oral administration of Seleno-Lentinan (SL2-1) markedly alleviated oxidative damage in the liver of mice induced by D-gal. In addition, SL2-1 significantly increased total antioxidant capacity, activities and protein expressions of catalase and glutathione peroxidase and lowered malondialdehyde levels in serum and liver. Fourier transform infrared spectroscopy analysis indicated that selenium of SL2-1 was mostly existed as the formations of OSeO, SeO and SeOC. Scanning electron microscope coupled with energy dispersive X-ray spectroscopy analysis revealed that the surface structure and elemental components of Lentinan significantly changed after selenizing. The results are instructive for the development of organic selenium-supplement resource.

Lentinan from shiitake mushroom (Lentinus edodes) suppresses expression of cytochrome P450 1A subfamily in the mouse liver.

Induction of xenobiotics metabolizing enzymes is related to the formation and chemoprevention of cancer. Since cytochrome P450s (CYPs) including CYP1A subfamily metabolize certain pro-carcinogens to their ultimate forms, down-regulation of CYP1As by food factors leads to the prevention of cancer. Mushroom polysaccharides, especially beta-glucans such as lentinan from Lentinus edodes, possess the anti-tumor and immunomodulating activities through the cytokine production from immunocytes. Recent our studies have demonstrated that lentinan suppresses hepatic CYP1As expression in the both constitutive and inducible levels through the production of tumor necrosis factor-alpha and an increase in the DNA-binding activity of nuclear factor-kappaB. This paper discusses on the effective lentinan dosage and route of administration for suppression of CYP1As.

Lentinan degradation in the Lentinula edodes fruiting body during postharvest preservation is reduced by downregulation of the exo-ß-1,3-glucanase EXG2.

Lentinan from Lentinula edodes fruiting bodies (shiitake mushrooms) is a valuable ß-glucan for medical purposes based on its anticancer activity and immunomodulating activity. However, lentinan content in fruiting bodies decreases after harvesting and storage due to an increase in glucanase activity. In this study, we downregulated the expression of an exo-ß-1,3-glucanase, exg2, in L. edodes using RNA interference. In the wild-type strain, ß-1,3-glucanase activity in fruiting bodies remarkably increased after harvesting, and 41.7% of the lentinan content was lost after 4 days of preservation. The EXG2 downregulated strain showed significantly lower lentinan degrading activity (60-70% of the wild-type strain) in the fruiting bodies 2-4 days after harvesting. The lentinan content of fresh fruiting bodies was similar in the wild-type and EXG2 downregulated strains, but in the downregulated strain, only 25.4% of the lentinan was lost after 4 days, indicating that downregulation of EXG2 enables keeping the lentinan content high longer.

Dynamic high pressure microfluidization-assisted extraction and antioxidant activities of lentinan.

Dynamic high pressure microfluidization (DHPM) was applied to assist the lentinan extraction. Response surface methodology (RSM), based on Box-Behnken design, was employed to optimize the DHPM-assisted extraction conditions of lentinan. Three main independent variables (DHPM pressure, ratio of water to raw material, extraction temperature) were taken into consideration. A yield of 7.200% was obtained under a modified condition (ratio of water to raw material of 65 mL/g, DHPM pressure of 147 MPa, extraction temperature of 83 °C), which matched well with the predicted value of the model. The molecular weight of the DHPM-assisted extract and hot water extract was 913,329 and 965,361 Da, respectively. Compared to the traditional hot water extraction, the lentinan extracted by DHPM assisting had better scavenging capacity of hydroxyl radical, superoxide anion free radical, 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical and nitrite. It could be concluded that the DHPM was a promising method to enhance the yield and antioxidant activity of lentinan during extraction.

Lentinus edodes: a macrofungus with pharmacological activities.

Lentinus edodes is the first medicinal macrofungus to enter the realm of modern biotechnology. It is the second most popular edible mushroom in the global market which is attributed not only to its nutritional value but also to possible potential for therapeutic applications. Lentinus edodes is used medicinally for diseases involving depressed immune function (including AIDS), cancer, environmental allergies, fungal infection, frequent flu and colds, bronchial inflammation, heart disease, hyperlipidemia (including high blood cholesterol), hypertension, infectious disease, diabetes, hepatitis and regulating urinary inconsistencies. It is the source of several well-studied preparations with proven pharmacological properties, especially the polysaccharide lentinan, eritadenine, shiitake mushroom mycelium, and culture media extracts (LEM, LAP and KS-2). Antibiotic, anti-carcinogenic and antiviral compounds have been isolated intracellularly (fruiting body and mycelia) and extracellularly (culture media). Some of these substances were lentinan, lectins and eritadenine. The aim of this review is to discuss the therapeutic applications of this macrofungus. The potential of this macrofungus is unquestionable in the most important areas of applied biotechnology.

Immunological effects of yeast- and mushroom-derived beta-glucans.

Glucans have a long history as nonspecific biological modulators. We compared the effects of three different glucans on immune reactions. Using two different administrations (intraperitoneal and oral) and two different animal models, we showed that yeast-derived Betamune (Biorigin, São Paulo, Brazil) caused significant stimulation of phagocytic activity as well as potentiation of synthesis and release of interleukin (IL)-1, IL-2, IL-4, IL-6, IL-8, IL-13, and tumor necrosis factor-alpha. In addition, Betamune inhibited growth of tumor cells in vivo and affected expression of several important genes in breast cancer cells. Compared to adult mice, young animals showed different sensitivity to glucan action.

Effects of excluded volume and polydispersity on solution properties of lentinan in 0.1 M NaOH solution.

Seven lentinan fractions of various weight-average molecular weights (M(w)), ranging from 1.45 x 10(5) to 1.13 x 10(6) g mol(-1) were investigated by static light scattering and viscometry in 0.1M NaOH solution at 25 degrees C. The intrinsic viscosity [eta] - M(w) and radius of gyration s(2)(z) (1/2) - M(w) relationships for lentinan in 0.1M NaOH solution were found to be represented by [eta] = 5.1 x 10(-3)M(w) (0.81) cm(3) g(-1) and s(2)(z) (1/2) = 2.3 x 10(-1)M(w) (0.58) nm, respectively. Focusing on the effects of the M(w) polydispersity with the Schulz-Zimm distribution function, the data of M(w), s(2)(z) (1/2), and [eta] was analyzed on the basis of the Yoshizaki-Nitta-Yamakawa theory for the unperturbed helical wormlike chain combined with the quasi-two-parameter (QTP) theory for excluded-volume effects. The persistence length, molecular weight per unit contour length, and the excluded-volume strength were determined roughly to be 6.2 nm, 980 nm(-1), and 0.1, respectively. Compared with the theoretical value calculated by the Monte Carlo model, the persistence length is longer than that of the single (1 --> 3)-beta-(D)-glucan chain. The results revealed that lentinan exists as single-stranded flexible chains in 0.1M NaOH solution with a certain degree of expansion due to the electrostatic repulsion from the interaction between the OH(-) anions and lentinan molecules.

Collapse and association of denatured lentinan in water/dimethlysulfoxide solutions.

Triple helical lentinan, beta-(1-->3)-D-glucan from Lentinus edodes, was denatured in dimethlysulfoxide (DMSO) into single random coils. The DMSO solutions of randomly coiled lentinan were diluted with pure water to different wH (the weight fraction of water in the mixed solvent), and their specific optical rotation [alpha]D, reduced viscosity (lnetar)/c, and hydrodynamic radius Rh were investigated as a function of wH and storage time t. With an increase of wH from 0.1 to 0.2, [alpha]D increased sharply, suggesting that transition of conformation of the macromolecules has occurred. When wH was lower than 0.1, (lnetar)/c of lentinan in water-diluted DMSO exhibited the almost same value as that in pure DMSO and changed hardly with increasing t. Interestingly, (lnetar)/c decreased to reach a minimum with a further increase of wH from 0.1 to 0.25 and then increased with a continuous increase of wH from 0.25 to 0.5. Both (lnetar)/c and Rh of the denatured lentinan in water-diluted DMSO with wH of approximately 0.25 both exhibited a minimum, indicating that collapsed coil chains have occurred. All of the experimental findings revealed that the behaviors of lentinan in water-diluted DMSO solution with wH < 0.1 were consistent with that in good solvent, DMSO. When wH = 0.25, the quality of the mixed solvents became worse, and the dominant intramacromolecular hydrogen-bond interaction enhanced, leading to minimum of viscosity and size of the chains as a result of the collapsed coils. When wH > 0.25, the quality of the mixture weakens further, and the intermolecular hydrogen-bond interaction enhanced and was dominant, leading to aggregation of the collapsed chains.