Fruit Bodies Of Grifola Frondosa Research Articles

Content of Extractive Substances and Polysaccharides in Fruit Bodies of Grifola frondosa Cultivated on Lignocellulose Substrates Depending on Extraction Methods

Content of Extractive Substances and Polysaccharides in Fruit Bodies of Grifola frondosa Cultivated on Lignocellulose Substrates Depending on Extraction Methods

A β-glucan from Grifola frondosa effectively delivers therapeutic oligonucleotide into cells via dectin-1 receptor and attenuates TNFα gene expression

Grifola frondosa is an edible and medicinal mushroom with great nutritional values and bioactivities. In the present study, a soluble homogeneous β-glucan, GFPS, with high molecular mass of 5.42 × 106 Da was purified from the fruit bodies of Grifola frondosa using 5% cold NaOH. The structure of GFPS was determined with FT-IR, NMR, and monosaccharide composition analysis, and was identified to be a β-D-(1-3)-linked glucan backbone with a single β-D-(1-6)-linked glucopyranosyl residue branched at C-6 on every third residue. Our results indicated that GFPS had a triple helical structure and could form complex with polydeoxyadenylic acid (poly[A]). Further studies demonstrated that GFPS could interact with poly[A] moiety of a designed antisense oligonucleotide (ASO) targeting the primary transcript of proinflammatory cytokine TNFα (TNFα-A60). This GFPS-based complex could incorporate TNFα-A60 into the macrophage cells via dectin-1 receptor and attenuate lipopolysaccharide-induced secretion of TNFα. Our results suggested that GFPS could be applied to deliver therapeutic oligonucleotides for the treatment of diseases such as inflammation and cancers.

Purification, characterization and immunomodulatory activity of a novel polysaccharide from Grifola frondosa

A novel bioactive polysaccharide, GFP-22, was isolated from the fruit bodies of Grifola frondosa by anion-exchange and gel filtration chromatography. Structure of GFP-22 was investigated using gas chromatography–mass spectrometry (GC–MS), methylation, nuclear magnetic resonance (NMR), high-performance size exclusion chromatography-multi-angle laser light scattering-refractive index detector (HPSEC-MALLS-RI) and atomic force microscopy (AFM) analysis. The backbone of GFP-22 is composed of 1,4-β-d-Glcp, 1,3-β-d-Glcp, 1,6-α-d-Glcp, 1,6-α-d-Galp, 1,4,6-α-d-Manp and 1,3,6-α-d-Manp units. Molecular weight of GFP-22 is 2.72 × 104 Da. GFP-22 has a linear filamentous structure. The administration of GFP-22 could improve or reverse the CTX-induced immunosuppression, significantly enhance the spleen and thymus indices, spleen lymphocyte proliferation and cytokines production in splenocytes. These findings suggest that GFP-22 could be explored as a natural and potential immunomodulatory agent and as an alternative means of lessening chemotherapy-induced immunosuppression.

Inducement of Cytokine Release by GFPBW2, a Novel Polysaccharide from Fruit Bodies of Grifola frondosa, through Dectin-1 in Macrophages

Polysaccharides, especially β-glucans isolated from various species of mushrooms, are considered as biological response modifiers (BRMs) to be widely used in the treatment of cancer, especially due to their immunostimulatory activity. We herein characterized the structure of a novel water-soluble homogeneous polysaccharide (GFPBW2) from the fruit bodies of mushroom Grifola frondosa and investigated its immunomodulatory activity in vitro. GFPBW2 was purified from the alkali-extracted fractions by stepwise elution with a molecular weight of 26.2 kDa. On the basis of infrared and NMR spectroscopy, methylation and monosaccharide composition analysis, partial acid hydrolysis, and Smith degradation, its structure was elucidated to possess a backbone consisting of β-d-1,3- and β-d-1,4-linked glucopyranosyl residues, with branches attached to O-6 of β-d-1,3-linked glucopyranosyl residues. Functionally, it is an effective inducer of tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6) secretion in murine resident peritoneal macrophages. Using quartz crystal microbalance (QCM) analysis, we found that GFPBW2 could bind dendritic cell-associated C-type lectin-1 (Dectin-1) with an affinity constant (Kd) value of 1.08 × 10 (-7) M, while it could activate Syk and enhance TNF-α production in RAW264.7 cells overexpressing wild type but not mutant Dectin-1. Furthermore, Syk, NF-κB signaling, and cytokine release in resident peritoneal macrophages induced by GFPBW2 could be significantly inhibited by a specific Dectin-1 blocking reagent, Laminarin. These data suggested that GFPBW2 might be a potential ligand of Dectin-1, and the potential of GFPBW2 to activate macrophage through triggering cytokine secretion might be attributed, at least in part, to the involvement of Dectin-1.

A heteropolysaccharide, l-fuco-d-manno-1,6-α-d-galactan extracted from Grifola frondosa and antiangiogenic activity of its sulfated derivative

The tumor growth and metastasis are angiogenesis dependent, thus blockade of angiogenesis is a promising approach for treatment of cancer. Herein we reported the structural and biological features of a novel water-soluble polysaccharide named GFPW from the fruit body of Grifola frondosa. Chemical and spectral analysis revealed that GFPW with an average molecular weight of 15.7kDa, possessed a backbone consisting of α-1,6-linked galactopyranosyl residues, with branches attached to O-2 of α-1,3-linked fucose residues and α-terminal mannose. By the chlorosulfonic acid–pyridine method, we prepared a sulfated derivative of GFPW, Sul-GFPW, with a substitution degree of 0.33. According to the 13C NMR spectrum, the substitution position was deduced at C-2 and C-3. The angiogenesis assays in vitro showed that Sul-GFPW significantly inhibited endothelial cell proliferation in a dose- and time-dependent manner, and reduced endothelial cell migration and tube formation as well.

Application of the Enzymolysis and Membrane Technology in Glucan Extraction and Purification from <i>Grifola frondosa</i>

It has been investigated that the application of the enzymolysis and membrane technology in the extraction and purification of glucan from Grifola frondosa. The fruitbody of Grifola frondosa was been used to extract polysaccharides. Compared the yield, protein content and polysaccharide content in the extracts under 3 different process modes, it was found that the procedure processed by complex enzyme was better than the others. Microfiltrated with membrane of 0.2μm aperture and then ultrafiltrated with membrane with molecular weight cutoff of 1000 Dalton, it had glucan retained rate of 80.42% and protein removed rate of 88.27%. GCP experiment indicated that the glucan get from the procedure was a homogeneous polysaccharide with weight average molecular weight of 130355 Dalton. The index of molecular weight distribution was 1.1844. The desalination rate was increased by adding water to the concentrate while ultrafiltration was in progress. When the quantity of water added was 1.5 times of the concentrate volume, it had effective desalination effect with the ash content of 3.79% in final product.

Structure of a β-glucan from Grifola frondosa and its antitumor effect by activating Dectin-1/Syk/NF-κB signaling

A soluble homogeneous β-glucan, GFPBW1, with a molecular mass of 300 kDa was purified from the fraction of the fruit bodies of Grifola frondosa extracted with 5% NaOH. Using various methods, such as infrared spectroscopy, NMR, methylation and monosaccharide composition analysis, its structure was determined to be a β-D-(1-3)-linked glucan backbone with a single β-D-(1-6)-linked glucopyranosyl residue branched at C-6 on every third residue. It induced TNF-α and IL-6 production and the activation of Syk and NF-κB signaling in resident peritoneal macrophages from ICR mice, which could be significantly inhibited by the blocking reagent laminarin. A competitive phagocytosis assay with FITC-zymosan indicated that GFPBW1 could bind to DC-associated C-type lectin 1 (Dectin-1). The TNF-α secretion and activation of Syk/NF-κB signaling triggered by GFPBW1 were enhanced in RAW264.7 cells overexpressing wild but not mutant (Δ38 and Y15S) Dectin-1. Furthermore, GFPBW1 potentiated the Concanavalin A-induced proliferative response of splenocytes and inhibited Sarcoma-180 growth allografted in ICR mice but not in immunodeficient BALB/c nu/nu mice. These results suggested that the antitumor activity of GFPBW1 was partially associated with the activation of macrophages via the Dectin-1/Syk/NF-κB signaling pathway. This molecule could be a promising biological response modifier with clear application for antitumor therapies.

Maitake beta-glucan enhances granulopoiesis and mobilization of granulocytes by increasing G-CSF production and modulating CXCR4/SDF-1 expression

Previous studies have presented that Maitake beta-glucan (MD-Fraction) extracted from the fruit body of Grifola frondosa has an anti-tumor effect by activating the immune system. Recently, the stimulating effects of beta-glucans on hematopoiesis were identified as new characteristics of polysaccharides, possibly helping to relieve the immunosuppression which results from chemotherapies. We demonstrated that the production of granulocyte colony-stimulating factor (G-CSF) was significantly enhanced by MD-Fraction (8 mg/kg, i.p.) in granulocytopenic model induced in mice using cyclophosphamide (200 mg/kg, i.p.). In addition, MD-Fraction induced a biphasic increase in the number of granulocytes in the spleen. The mechanism for the increase in granulocytes on the early phase on day 1 might involve the increased mRNA expression of macrophage inflammatory protein-2 (MIP-2), in the splenic cells, thereby recruiting granulocytes into the spleen. Interestingly, a decline of myeloid progenitors in the bone marrow and an increase in granulocytes in the peripheral blood were observed on day 5, suggesting a mobilization of granulocytes and their progenitors from the bone marrow to the peripheral blood. We confirmed that a possible mechanism in which MD-Fraction promoted the mobilization of granulocytes and their progenitors from the bone marrow is down-regulating the expression of the chemokine receptor, CXCR4, and its ligand, stromal cell-derived factor 1 (SDF-1) in the bone marrow microenvironment. These results reveal a novel function of Maitake beta-glucan that enhances the granulopoiesis and mobilization of granulocytes and their progenitors by stimulating G-CSF production. This finding presents opportunities to develop new therapeutic strategies against the immunosuppression caused by chemotherapies in cancer patients.

이산화탄소가 잎새버섯의 자실체 발생 및 생육에 미치는 영향

Effects of concentration (500, 800, 1,000 and 1,500 ppm) on the initiation and development of fruit body of Grifola frondosa on sawdust cultivation were studied. Optimum concentrations of carbon dioxide for the initiation and development of the fruit body showed the ranges from 500 to 800 ppm. Fruit body initiation was accelerated at lower than 800 ppm exposure but the maturing of the fruit body was not influenced by above treated concentrations. The higher ratio of primordial formation, faster fruit body initiation and higher yield were obtained at below 800 ppm of level, whereas over 1,000 ppm of levels showed abnormal and lower quality of fruiting bodies. Based on the above results, it is concluded that the favorable level for bag culture of G. frondosa was below 800 ppm.

Inhibitory Effect of TNF-α Produced by Macrophages Stimulated withGrifola frondosaExtract (ME) on the Growth of Influenza A/Aichi/2/68 Virus in MDCK Cells

We investigated the inhibitory effect of the conditioned medium (CM) from P338D1 (D1) cells, a murine macrophage cell line, stimulated for 10 hours with a fixed dose (100 mug/ml) of the extracts from the fruit bodies of Grifola frondosa (ME) or its ultra filtration-based fractions (MFs), on the growth of influenza A/Aichi/2/68 virus in Madin-Darby canine kidney cells. Direct addition of ME and 3 kinds of MFs (MF1, MF2 and MF3) to the infected cells had no obvious inhibitory effect. However, virus yields were reduced in the presence of CMs. Notably, the inhibitory effect of the CM prepared by using MF2 (molecular weight of 30 Kd to 100 Kd) was the strongest (28% reduction compared to the control). RT-PCR and ELISA assays showed that the CMs could induce the expression of TNF-alpha mRNA in D1 cells leading to production of TNF-alpha, known as an antiviral cytokine. These findings suggest that ME and MFs (especially MF-2) might induce the production of certain factors, including TNF-alpha, which are responsible for the inhibition of viral growth in vitro.

An Optical Measurement Method Employing Voltage-sensitive Dyes for Investigation of the Potential Spread Inside the Fruit Body of Grifola frondosa

本論文では, 子実体の生体電位の微視的な発生および伝播のダイナミクスの解明を目的とし, 膜電位感受性色素を用いた光学的計測法による, 細胞レベルでの電位変化の計測を試みた. その結果, 波長540 nm ではパルス電流刺激により吸光度は一時的に増大し, 波長700 nm では吸光度は一時的に減少するといった, 膜電位感受性色素由来の吸光度の反転現象｢波長反転｣を確認することができた. これより, 計測で得た光学的変化が菌糸細胞の電位変化に伴うものであると結論した. また, 刺激に対する電位反応が菌糸細胞上を伝播する様子を明らかにするとともに, これまでに実施してきた光刺激時の子実体の生体電位応答特性について, その発生ダイナミクスに関する考察を得た. これらの結果を踏まえ, 今後はこれまでに報告してきた各種外的刺激に対する子実体の生体電位反応過程について, 電気回路によるモデル化を試みる予定である.

Alpha-glucosidase inhibitor from the seeds of balsam pear (Momordica charantia) and the fruit bodies of Grifola frondosa.

Alpha-glucosidase inhibitory activities were found in aqueous methanol extracts of the seeds of Momordica charantia and the fruit bodies of Grifola frondosa. An active principle against the enzyme prepared from rat small intestine acetone powders was isolated and characterized. The structure of the isolated compound was identified as D-(+)-trehalose by FDMS, 1H-, 13C-NMR, and [alpha]D measurements. The inhibitory activity of trehalose was compared with 1-deoxynojirimycin. Trehalose showed 45% inhibitory activity at the concentration of 2 x 10(-3) m, but 1-deoxynojirimycin had 52% inhibitory activity at 1 x 10(-7) M.

ChemInform Abstract: Constituents of Mushrooms. Part 9. Structures of New Ceramides from the Fruit Bodies of Grifola frondosa.

AbstractChemInform is a weekly Abstracting Service, delivering concise information at a glance that was extracted from about 100 leading journals. To access a ChemInform Abstract of an article which was published elsewhere, please select a “Full Text” option. The original article is trackable via the “References” option.

Structures of new ceramides from the fruit bodies of Grifola frondosa.

Four new phytosphingosine-type ceramides, (2S,3S,4R)-2-[(2'R)-2'-hydroxydocosanoylamino]-1,3,4-octadecane triol (1), (2S,3S,4R)-2-[(2'R)-2'-hydroxytricosanoylamino]-1,3,4-octadecan etriol (2), (2S,3S,4R)-2-[(2'R)-2'-hydroxypentacosanoylamino]-1,3,4-octadec anetriol (3) and (2S,3S,4R)-2-[(2'R)-2'-hydroxyhexacosanoylamino]-1,3,4-octadeca netriol (4), have been isolated from the fruit bodies of Grifola frondosa. The structures of the new compounds were elucidated on the basis of their spectral data.

ChemInform Abstract: Sterol Constituents from the Fruit Bodies of Grifola frondosa (FR.) S. F. GRAY

AbstractChemInform is a weekly Abstracting Service, delivering concise information at a glance that was extracted from about 100 leading journals. To access a ChemInform Abstract of an article which was published elsewhere, please select a “Full Text” option. The original article is trackable via the “References” option.

Sterol Constituents from the Fruit Bodies of Grifola frondosa (FR.) S. F. GRAY.

Four new sterols, 5α, 6α-epoxy-(22E, 24R)-ergosta-8(14), 22-diene-3β, 7β-diol (1), (22E, 24R)-ergosta-8, 22-diene-3β, 5α, 6β, 7α-tetrol (2), (22E, 24R)-ergosta-7, 9(11), 22-triene-3β, 5α, 6β-triol (3) and 3β, 5α, 6β-trihydroxy-(22E, 24R)-ergost-22-en-7-one (4), have been isolated from the fruit bodies of Grifola frondosa (FR.) S. F. GRAY (Polyporaceae)together with fourteen known ones (5-18), of which two (5 and 6) are reported for the first time from a natural source. The structures of these compounds were elucidated on the basis of spectral data.

Anti-diabetic activity present in the fruit body of Grifola frondosa (Maitake). I.

The fruit body of Grifola frondosa (maitake), Basidiomycetes was confirmed to contain substances with anti-diabetic activity. When 1 g/d of powdered fruit body of maitake was given orally to a genetically diabetic mouse (KK-Ay), blood glucose reduction was observed, in contrast to the control group in which the blood glucose increased with ageing. Moreover, levels of insulin and triglyceride in plasma demonstrated a change similar to blood glucose with feeding of maitake. Ether-ethanol-soluble (ES) and hot water-soluble (WS) fractions were prepared from the fruit body and their hypoglycemic activity was examined. Blood glucose-lowering activity was found when ES-fraction or WS-50% ethanol float (X) fraction was administered orally, but other WS-fractions were inactive. These results suggest that the anti-diabetic activity was present not only in the ES-fraction consisting of lipid but also in the X-fraction of peptidoglycan (sugar:protein = 65:35).

Antitumor activity exhibited by orally administered extract from fruit body of Grifola frondosa (maitake).

The acid-insoluble, alkali-soluble, hot-water-extractable polymer (a polysaccharide containing approximately 30% of protein; D-fraction) obtained from the fruit bodies of Grifola frondosa (maitake) exhibited antitumor activities against allogenic and syngeneic tumors on oral administration to mice. Winn assay conducted to examine the tumor growth-suppressing effect revealed a complete inhibition of the tumor by the oral administration of D-fraction. This fact indicates that stimulation of the immune response system triggered by the tumor-bearing state is activated by D-fraction. Consequently, the activity of D-fraction on cells associated with immune response was examined. The cytolytic activity and interleukin-1 productivity of macrophages or T cells which exhibit antigen-specific cytotoxicity were enhanced. D-fraction was found to potentiate the delayed-type hypersensitivity response which is associated with tumor growth suppression.

Blood pressure-lowering activity present in the fruit body of Grifola frondosa (maitake). I.

The fruit body of Grifola frondosa (maitake), Basidiomycetes, was confirmed to contain a substance with blood pressure-lowering activity. When powdered fruit body of maitake was given orally to spontaneously hypertensive rats (SHR), blood pressure reduction was observed, in contrast to the control group in which the blood pressure increased with ageing. Ether-soluble (ES) and hot water-soluble (WS) fractions were prepared from the fruit body and their hypertensive action was examined. Blood pressure-lowering activity was found when the dried ES fraction was administered orally at 30mg/kg, but the WS fraction was inactive. Thus, the ES fraction was further separated into setone-soluble (ES-AS) and insoluble (ES-AP) fractions. Administration of 20mg/kg dried material caused a blood pressure reduction of about 45mmHg in the former case and 65mmHg in the latter as compared with the control (about 220mmHg). When the EA-AP fraction was separated to 5 subfractions by thin layer chromatographic analysis, the most potent activity was observed in the subfraction which was detected on the spot of Rf=55.3. When authentic antihypertensive agents were examined in combination with maitake, a hypotensive effect was observed on administration of reserpine. This suggests that the blood pressure-lowering effect of maitake was brought about by a mechanism other than sympatholytic action.

The chemical structure of an antitumor polysaccharide in fruit bodies of Grifola frondosa (maitake).

A polysaccharide was extracted from fruit bodies of Grifola frondosa (maitake), and the chemical structure and antitumor activity were studied. The extracted polysaccharide could be hydrolyzed by ?A-glucanase into glucose, indicating it to be a β-glucan. The sample gave methyl 2, 3, 4, 6- tetra-Ο-, methyl 2, 4, 646-Ο-, methyl 2, 3, 4-tri -Ο- and methyl 2, 4-di-Ο-methylglucoside in the molar ratio of 4 : 2 : 1 : 4 on methylation. In the carbon-13 nuclear magnetic resonance spectrum, the signals of C-6' (related to (1-6) bonding) and C-3' (related to (1-3) bonding) were observed in addition to those of free C-6 and C-3. These results indicate that the major chain is made up of β-1, 6-linked glucose residues with branches of β-1, 3-linked glucose. This glucan inhibited the growth of Sarcoma 180 tumor in ICR mice.