ofGanoderma Lucidum Research Articles

Resource Utilization Residue of <i>Pleurotus ostreatus</i>

The allelopathy of spent substrate extracts including water extract and ethanol extract ofPleurotus ostreatuson the mycelium growth of six edible fungi, i.e.Flammulina velutipes,Ganoderma lucidum Karst,Pleurotus ostreatusandCordyceps (three species), were investigated using Petri dishes approach. The results indicated that the spent substrate extracts have different effects on their mycelium growth. The mycelium ofFlammulina velutipesgrows better than control check with increasing water extract concentration. The mycelia ofGanoderma lucidum Karstgrow first promotion after inhibition with increasing water extract concentration. The mycelia ofFlammulina velutipes, Ganoderma lucidum Karst, Pleurotus ostreatusare promoted by ethanol extracts. The mycelia of Cordyceps (three species) grow first promotion after inhibition with ethanol extracts. The results can provide reference values for rational utilization of the spent mushroom substrate.

Synergistic Cytotoxic Effects of Ganoderma lucidum and Bacillus Calmette Guérin on Premalignant Urothelial HUC-PC Cells and Its Regulation on Proinflammatory Cytokine Secretion.

Bacillus Calmette-Guérin (BCG) is conventionally used as an adjuvant immunotherapy to reduce the recurrence of bladder cancer. To address the issues of efficacy and safety, an ethanol extract of Ganoderma lucidum (GLe) was evaluated for its interaction with BCG. In a model of premalignant human uroepithelial cells (HUC-PC), GLe exerted immediate cytotoxic effects while BCG showed a delayed response, given that both were immunological active in inducing the secretion of interleukin (IL)-6, IL-8, and monocyte chemotactic protein-1 (MCP-1). Synergistic cytotoxic effects were observed when cells were either coincubated with both drugs or firstly preincubated with GLe. Synergism between GLe and BCG was demonstrated to achieve a complete cytostasis in 24 hours, and such effects were progressed in the subsequent 5 days. However, the pretreatment of GLe resulted in suppression of IL-6, IL-8, and MCP-1 secretions without affecting the cytotoxicity. Given that numerous proinflammatory cytokines are associated with the high side effects toll of BCG, results herein suggested the potential implications of GL to supplement the BCG immunotherapy in bladder cancer, for better efficacy and reducing side effects.

Structural Characterization and Antioxidative Activity of Low-Molecular-Weights Beta-1,3-Glucan from the Residue of ExtractedGanoderma lucidumFruiting Bodies

The major cell wall constituent of Ganoderma lucidum (G. lucidum) is β-1,3-glucan. This study examined the polysaccharide from the residues of alkaline-extracted fruiting bodies using high-performance anion-exchange chromatography (HPAEC), and it employed nuclear magnetic resonance (NMR) and mass spectrometry (MS) to confirm the structures. We have successfully isolated low-molecular-weight β-1,3-glucan (LMG), in high yields, from the waste residue of extracted fruiting bodies of G. lucidum. The 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay evaluated the capability of LMG to suppress H2O2-induced cell death in RAW264.7 cells, identifying that LMG protected cells from H2O2-induced damage. LMG treatment decreased H2O2-induced intracellular reactive oxygen species (ROS) production. LMG also influenced sphingomyelinase (SMase) activity, stimulated by cell death to induce ceramide formation, and then increase cell ROS production. Estimation of the activities of neutral and acid SMases in vitro showed that LMG suppressed the activities of both neutral and acid SMases in a concentration-dependent manner. These results suggest that LMG, a water-soluble β-1,3-glucan recycled from extracted residue of G. lucidum, possesses antioxidant capability against H2O2-induced cell death by attenuating intracellular ROS and inhibiting SMase activity.

Antiperoxidative, anti-inflammatory, and antimutagenic activities of ethanol extract of the mycelium of Ganoderma lucidum occurring in South India.

Free radical mediated genetic instability is widely thought to be a major etiological factor for initiation of carcinogenesis. Mushrooms represent a largely untapped source of powerful new pharmaceutical products. In the present study, we examined the antiperoxidative, anti-inflammatory, and antimutagenic activities of the ethanol extract of the mycelium of a medicinal mushroom, Ganoderma lucidum, occurring in south India. Antiperoxidative activity was evaluated using Fe(2+)-ascorbate-induced lipid peroxidation in rat liver homogenate and a phorbol ester (croton oil)-induced lipid peroxidation in mouse skin. Antiinflammatory activity was evaluated against carrageenan-induced acute and formalin-induced chronic inflammatory paw edema in mouse and phorbol ester-induced mouse skin inflammation. Antimutagenic activity was determined by the Ames mutagenicity assay using histidine mutant of Salmonella typhimurium strains TA 98, TA100, and TA102. Sodium azide (NaN(3)), N-methyl-N-nitro-N-nitrosoguanidine (MNNG), 4-nitro-o-phenylenediamine (NPD), and benzo[a]pyrene (B[a]P) were used as the mutagens. The extract showed significant inhibition of Fe(2+)-induced peroxidation of lipid in rat liver (IC(50) 510 +/- 22 microg/ml) and 37% inhibition of croton oil-induced peroxidation on the mouse skin at 20 mg/0.1 ml/skin. Carrageenan-induced acute and formalin-induced chronic inflammatory edema were inhibited by 56 and 60%, respectively, by the extract at 1,000 mg/kg body wt (i.p). The extract at a concentration of 5 mg/plate showed inhibition of mutagenicity elicited by direct acting mutagens, NaN(3) (55.5 and 75.7%) and MNNG (50.0 and 57.5%) for S. typhymurium strains TA100 and TA102, respectively. The extract at the same concentration also inhibited mutagenicity elicited by NPD (52.4 and 64.2%) and B[a]P (60.7 and 59.6%) for TA98 and TA100 strains, respectively. The B[a]P was activated in the presence of rat liver microsomal (S9) fraction. The results of our study revealed that ethanol extract of Ganoderma lucidum mycelium possessed significant antiperoxidative, antiinflammatory, and antimutagenic activities. The findings suggest a medicinal use for the ethanol extract of the mycelium of G. lucidum occurring in South India.

Growth and fruitbody formation of Ganoderma lucidum on media supplemented with vanadium, selenium and germanium

Responses of mycelia ofGanoderma lucidum to vanadium, selenium and germanium were examined over a wide range of concentrations (10–1, 120 μg/ml) in pure culture. Se and V were found to be highly toxic, but Ge was not toxic at the levels tested.Ganododerma lucidum cultivated on substrates of sawdust with V (30–80 μg/g) developed mature fruitbodies, but the bioaccumulation of V was quite low (2.5–7 μg/g in pileus, 12.5–21.5 μg/g in stipe and <1 μg/g in basidiospores). Se as Na2SeO4 labeled with75Se was effectively taken up from substrates and accumulated in fruitbodies (mainly in pileus), then depleted by discharge of basidiospores. Ge as GeCl4 labeled with77Ge was easily uptaken and translocated into fruitbodies.

Inhibition of cytopathic effect of human immunodeficiency virus-1 by water-soluble extract ofGanoderma lucidum

To examine components ofGanoderma lucidum for anti-human immunodeficiency virus (HIV) activity, the aqueous extracts of its basidiocarps were separated into high-molecular-weight (HMF) and low-molecular-weight (LMF) fractions. These fractions were used in XTT [2,3-bis (2-methoxy-4-nitro-5-sulfophenyl)-5-[(phenylamino) carbonyl]-2H-tetrazolium hydroxide] antiviral assay which can quantitatively measure cytopathic effects of HIV-1 on CEM, human T lymphoblastoid cell line. The CEM cell line added with serial diluted HMF or LMF was cultured in the absence or presence of HIV-1. The results showed that the LMF of the aqueous extract strongly inhibited cytopathic effect of the target cell induced by HIV-1. When two-fold serially diluted LMF ranging from 0.97 mug/ml to 125.00 mug/ml was added to the virus-free culture system, no toxicity on the target cells was detected in all the concentrations tested. However, when it was added to the HIV-infected culture system, the viabilities of the target cell reached a plateau recovering its viabilities to 71.7% and 82.5% in experiment-1 and-2 at 15.60 mug/ml, respectively. The cell viabilities were then gradually decreased but maintained at more than 50% above 31.20 mug/ml concentration. On the contrary, HMF did not prevent any HIV-induced cytopathic effect at any concentrations tested on this cell line. From these results, negligible toxicities were observed by both HMF and LMF ofG. lucidum, and recovery of cell viability in HIV infected target cell was induced only by LMF of the carpophores.

Anti-Helicobacter pylori activity of mushrooms

Inhibitory effects of the mushrooms on the growth and urease ofHelicobacter pylori (HP), which is associated with human gastroduodenal diseases such as gastritis, peptic ulcer and gastric carcinoma, were investigated. Most of the mushroom extracts did not show inhibitory effect on HP urease exceptCoriolus versicolar, Auricularia auricula, Sarcodon aspratus andFlammulina velutipes. The extract ofGanoderma lucidum, Coriolus versicolar, Gyropora esculenta andAgaricus bisporus var. albidus inhibited the growth of HP. When their extracts were fractionated, the ether fraction ofGanoderma lucidum andAgaricus bisporus var. albidus were the most effective. Among seven components seperated from the ether fraction ofG. lucidum extract by silica gel column chromatography, P3 was the most potent: MIC was 200 μg/ml. However, P3 did not inhibit the urease.

Enhancement of water-solubilities of protein-bound polysaccharides contained in the basidiocarps ofGanoderma lucidum by hydrolyzing with chymotrypsin

Optimum conditions for hydrolysis were investigated to enhance water-solubilities of protein-bound polysaccharides in the basidiocarps ofGanoderma lucidum by treating chymotrypsin. We also attempted withGanoderma lucidum residue remaining after extracting hot water-soluble components inGanoderma lucidum. After hydrolyzing under optimum conditions (20 ppm chymotrypsin, 2%Ganoderma lucidum or 6%Ganoderma lucidum residue, at pH 10 and at 40°C), the amounts of total protein and carbohydrate of hydrolysate were measured. Michaelis constant, Km and maximum rate, Vmax calculated by Lineweaver-Burk plot for the hydrolysis ofGanoderma lucidum were 1.73% and 0.073%/min respectively and those for hydrolysis ofGanoderma lucidum residue were 2.40% and 0.033%/min respectively. The amount of polysaccharide isolated fromGanoderma lucidum (100 g) treated with chymotrypsin was only 3.07 g, but significantly increased amount (14.34 g) of polysaccharides was isolated fromGanoderma lucidum residue (100 g) treated with chymotrypsin. The protein-bound polysaccharide was isolated from the non-hydrolyzed and hydrolyzed samlpe and molecular weights of the polysaccharide were measured by Sepharose CL-4B gel filtration.

Conversion of water-insoluble components of the basidiocarps ofGanoderma lucidum to water-soluble components by hydrolyzing with chitinase

We investigated the optimum conditions for conversion of water-insoluble components of basidiocarps ofGanoderma lucidum to water-soluble components by hydrolyzing with chitinase. We also tried it withGanoderma lucidum residue remaining after extracting hot water-soluble components ofGanoderma lucidum. After hydrolyzing under optimum conditions (20 ppm chitinase, 2%Ganoderma lucidum or 6%Ganoderma lucidum residue, at pH 3 and at 35°C), the contents of total water-soluble components (polysaccharide or protein) were measured, and it was found that the contents of water-soluble components increased to 1.5∼2.7 fold. Michaelis constant, Km and maximum rate, Vmax calculated by Lineweaver-Burk plot for hydrolysis ofGanoderma lucidum were 1.75% and 0.02%/min respectively and those for hydrolysis ofGanoderma lucidum residue were 53.15% and 0.53%/min respectively. The protein-bound polysaccharide was isolated after hydrolysis and molecular weights were measured by Sepharose CL-4B gel filtration and compared with the molecular weights of polysaccharide before hydrolysis.

Formation of atypical fruiting structures in Ganoderma lucidum isolates on a nutrient agar medium

Effects of light and ventilation on the formation of atypical fruiting structures (AFSs) and fruit body primordia (FBPs) ofGanoderma lucidum on nutrient agar media were investigated. Although the mycelial growth was inhibited by illumination and ventilation, brown AFSs appeared on the white mycelial colony, and basidia and basidiospores were produced on the AFSs. On the other hand, FBPs were induced by illumination alone, regardless of ventilation. However, the primordia could not develop to mature fruit bodies. In the dark, only vegetative growth of the fungus progressed. Twenty-three isolates ofG. lucidum collected from four countries were tested for the formation of AFSs and FBPs under light and ventilation. Thirteen isolates formed AFSs, and another five isolates produced FBPs. Of the remaining five isolates, one formed callus-like structures without elaborating basidiospores, and the other four did not induce AFSs or FBPs. Microscopical observation showed that the basidia were formed directly from generative hyphae on the surface of AFSs. Basidiospores formed on the basidia were brown and ellipsoid with an eccentric hilar appendix on the rounded spore base. They had a double wall and mostly contained one or two large vacuoles. The surface of basidiospores was smooth or wrinkled and had shallow holes. The spore size was (4.5−)6.4−9.6(−10.3) × (2.6−)3.2−5.1(−6.4), 7.3 × 4.2 µm on average.

Antimicrobial activity of Ganoderma lucidum extract alone and in combination with some antibiotics.

Antimicrobial activity of GL (the aqueous extract from the carpophores of Ganoderma lucidum (FR)KARST) was tested in vitro against Gram positive and Gram negative bacteria by serial broth dilution method, and the antimicrobial activity was expressed by minimal inhibitory concentration (MIC). Among fifteen species of bacteria tested, the antimicrobial activity of GL was the most potent against Micrococcus luteus (MIC, 0.75 mg/ml). To investigate the effects of antimicrobial combinations of GL with four kinds of antibiotics (ampicillin, cefazolin, oxytetracycline and chloramphenicol), the fractional inhibitory concentration index (FICI) was determined by checkerboard assay for each strain. The antimicrobial combinations of GL with four antibiotics resulted in additive effect in most instances, synergism in two instances, and antagonism in two instances. Synergism was observed when GL was combined with cefazolin against Bacillus subtilis and Klebsiella oxytoca.

Anti-allergic constituents in the culture medium ofGanoderma lucidum. (I) Inhibitory effect of oleic acid on histamine release

The chloroform extract from Ganoderma lucidum broth markedly inhibited histamine release from rat peritoneal mast cells. From the active fractions, palmitic acid, stearic acid, oleic acid and linoleic acid were isolated. Oleic acid dose-dependently inhibited the histamine release and 45Ca uptake into mast cells induced by compound 48/80 and A-23187 at concentrations of 5 to 50 microM and 0.5 to 5 microM, respectively. Saturated fatty acids, however, had only a weak inhibitory effect on histamine release. Although linoleic acid and linolenic acid effectively prevented this release, these two compounds caused marked release at concentrations higher than 10 microM and 20 microM, respectively. Oleic acid induces membrane-stabilization in model membrane systems. It was concluded that one of the effective constituents obtainable from the chloroform extract of G. lucidum-cultured broth is oleic acid.

Anti-allergic constituents in the culture medium ofGanoderma lucidum. (II) The inhibitory effect of cyclooctasulfur on histamine release

For centuries, Ganoderma lucidum has been used in Oriental medicine for the treatment of chronic bronchitis. Sequential fractions of the culture medium of this plant revealed that one of the active constituents was cyclooctasulfur. The latter effectively inhibited histamine release from rat peritoneal mast cells and impeded 45Ca uptake into these cells without affecting the cyclic AMP content. SDS-PAGE analysis indicated that cyclooctasulfur induced some changes in protein bands obtained from the membrane fraction of mast cells, suggesting that this compound interacts with membrane proteins so as to inhibit 45Ca uptake, and that this may be the main cause of histamine release inhibition.

Studies on protoplast formation and regeneration ofGanoderma lucidum

To obtain a new strain ofGanoderma lucidum by protoplast fusion technique, its protoplast formation and regeneration were studied. Several factors affecting the protoplast formation and regeneration were investigated to find their optimum conditions. The mycelium was grown for four days on the cellophane membrane placed onG. lucidum complete medium (GCM). When various commercial lytic enzymes were examined for protoplast isolation, the combination of Novozym 234 and β-glucuronidase was found to be effective. An osmotic stabilizer, 0.6 M sucrose in 20 mM phosphate buffer pH 5.8, gave the highest yield of protoplasts. Three-hour incubation in shaking incubator was most suitable for releasing protoplasts. To increase the protoplast yield, pretreatment with 2-mercaptoethanol was carried out. The regeneration frequency in GCM containing 0.6 M MgSO4·7H2O was shown to be 0.66%.

Novel Triterpenoids from the Mycelial Mat at the Previous Stage of Fruiting ofGanoderma lucidum

Novel Triterpenoids from the Mycelial Mat at the Previous Stage of Fruiting of<i>Ganoderma lucidum</i>

Spore-forms in sporophores ofGanoderma lucidum (Leyss.) Karst

1. In this paper information regarding the various spore-forms ofGanoderma lucidum has been presented.