Traditional use of the tiger milk medicinal mushroom, Lignosus rhinocerus, to treat various illnesses has been recorded for > 4 centuries. Successful cultivation of L. rhinocerus using proprietary solid-state fermentation (SSF) technology by LiGNO Biotech has enabled large-scale production of L. rhinocerus sclerotia (termed L. rhinocerus TM02) and further investigations into its medicinal properties. Pharmacological activities of L. rhinocerus TM02, including its antioxidant, anti-inflammatory, anticancer, and immunomodulatory effects and the bioactive components responsible, have been validated by various scientific studies. In this study, we assessed the consistency of the bioactive components in 11 batches of L. rhinocerus TM02 produced over a 9-year period. The different batches of L. rhinocerus TM02 consisted of stable protein, polysaccharide, and glycoprotein contents, and all tested samples were comparable to the wild type. L. rhinocerus TM02 had greater protein, carbohydrate, and glycoprotein contents, which were mostly bioactive compared to another cultivar from a different cultivation technology (TM-UN). Together with previous scientific validations, L. rhinocerus TM02 produced using SSF cultivation is of optimal quality with high consistent bioactive contents, which can be an appropriate indicator for quality validation of the much sought-after medicinal mushroom, L. rhinocerus.
Cordyceps militaris is known for its curative properties. The present study was undertaken to evaluate the reduction of nitric oxide production by BV2 cells by the bioactive fraction of stroma powder of C. militaris, and to deduce the potential chemical components and pathways that may be responsible. The CE2 fraction from ethyl acetate extract did not exert any cytotoxic effects toward the BV2 cells at concentrations 0.1 to 100 μg/mL. The CE2 fraction also showed a significant (p < 0.05) reduction in nitric oxide production at 1-100 μg/mL. At 10 μg/mL, the CE2 fraction attenuated 85% of the NO production in BV2 cells. Further, the CE2 fraction (10 μg/mL) downregulated inflammatory genes, iNOS and COX-2, and upregulated anti-inflammatory genes, HO-1 and NQO-1. The CE2 fraction reduced NO production via activation of NRF2 and NF-κB transcriptions. The chemical constituents of the bioactive CE2 fraction were identified via GCMS. Eleven lipid components were identified including fatty acids, fatty acid esters, and sterols.
Dyslipidemia is the key precursor of atherosclerotic cardiovascular disease. The aim of this study was to investigate the lipid-modifying potential of organic solvent-partitioned extracts from fruiting bodies of Amauroderma rugosum in vitro using oleate-induced human hepatocellular liver carcinoma (HepG2) cells. Our results demonstrated that oleate-induced HepG2 cells treated with ethyl acetate (EA) extract greatly decreased intracellular and secreted total triglyceride (TG) and total cholesterol (TC) compared with other extracts. Further investigation of cellular expression of selected apolipoproteins also revealed that oleate-induced HepG2 cells treated with the EA extract best attenuated the apolipoprotein (Apo) profile by downregulating ApoB-100 and ApoE while upregulating ApoA1. Because both ApoB-100 and ApoE are key components of low-density lipoprotein (LDL) and very LDL (VLDL), which are recognized as "bad cholesterol," this result indicates that treatment with the EA extract inhibited LDL and VLDL production in oleate-induced HepG2 cells. On the other hand, increasing ApoA1 evidence shows antiatherogenic benefits to increasing ApoA1, the key component of high-density lipoprotein (HDL), particularly in relation to its role in promoting reverse cholesterol transport and preventing LDL oxidation; this indicates that the EA extract upregulated the production of HDL ("good cholesterol"). Hence, the EA extract is a good source of lipid-ameliorating agents in the management of dyslipidemia.
Neurodegenerative disease is defined as a deterioration of the nervous system in the intellectual and cognitive capabilities. Statistics show that more than 80-90 million individuals age 65 and above in 2050 may be affected by neurodegenerative conditions like Alzheimer's and Parkinson's disease. Studies have shown that out of 2000 different types of edible and/or medicinal mushrooms, only a few countable mushrooms have been selected until now for neurohealth activity. Hericium erinaceus is one of the well-established medicinal mushrooms for neuronal health. It has been documented for its regenerative capability in peripheral nerve. Another mushroom used as traditional medicine is Lignosus rhinocerotis, which has been used for various illnesses. It has been documented for its neurite outgrowth potential in PC12 cells. Based on the regenerative capabilities of both the mushrooms, priority was given to select them for our study. The aim of this study was to investigate the potential of H. erinaceus and L. rhinocerotis to stimulate neurite outgrowth in dissociated cells of brain, spinal cord, and retina from chick embryo when compared to brain derived neurotrophic factor (BDNF). Neurite outgrowth activity was confirmed by the immu-nofluorescence method in all tissue samples. Treatment with different concentrations of extracts resulted in neuronal differentiation and neuronal elongation. H. erinaceus extract at 50 µg/mL triggered neurite outgrowth at 20.47%, 22.47%, and 21.70% in brain, spinal cord, and retinal cells. L. rhinocerotis sclerotium extract at 50 µg/mL induced maximum neurite outgrowth of 20.77% and 24.73% in brain and spinal cord, whereas 20.77% of neurite outgrowth was observed in retinal cells at 25 µg/mL, respectively.
Alzheimer's disease (AD) is the leading neurodegenerative disorder affecting memory and learning of aged people. Hypercholesterolemia had been implicated as one of the stark hallmarks of AD. Recent AD control guidelines have suggested lifestyle modification to slow down the progression of AD. In this regard, medicinal mushroom Ganoderma lucidum seems apt. In the present study, hot water extract of G. lucidum (200 mg/kg body weight) was fed to the hypercholesterolemic and AD model rats for 8 weeks. Nonspatial memory and learning abilities of the model animals was assessed using novel object recognition (NOR) test, rotarod test, and locomotor/open-field test. Then, the animals were sacrificed and transmission electron micrograph (TEM) view of the hippocampal neurons was assessed. In all the nonspatial memory and learning tests, the G. lucidum HWE fed rats performed better indicating improved memory and learning abilities. TEM view showed regular arrangement of the neurons in the G. lucidum HWE fed rats compared with those of the deranged arrangement of the AD rats. G. lucidum might have aided in restoring the memory and learning abilities of the AD model animals through maintaining neuronal structure and function. Thus, G. lucidum could be suggested as a medicotherapeutic agent against AD.
Hypercholesterolemia has been implicated as one of the pathomechanistic factors of Alzheimer's disease (AD), the most common neurodegenerative disorder affecting memory and learning abilities. In the present study, ameliorative effect of hot water extract (HWE) of mushroom Ganoderma lucidum to the memory and learning related behavioral performance of hypercholesterolemic and AD rats was investigated using Morris water maze (MWM). Male Wistar rats were randomly grouped into control, extract fed control, hypercholesterolemic, extract fed hypercholesterolemic, AD, and extract fed AD groups, each group containing 8 animals. Hypercholesterolemia was induced in rats by adding 1% cholesterol and 1% cholic acid with the basal diet of the respective group. Alzheimer's disease model rats were prepared through infusion of amyloid β(1-42) to the right ventricle. Memory and learning related performance of all the rats was tested for 6 consecutive days that included time taken to reach the submerged platform (sec) and distance traveled (m). G. lucidum HWE fed rats took less time and traveled less distance to find the submerged platform, which indicates the spatial learning and memory related behavioral amelioration of the extract fed rats compared with their non-fed counterparts. Thus, usage of G. lucidum seems promising in withstanding hypercholesterolemia-induced Alzheimer's disease pathogenesis.
The growth and lectin production of Ganoderma applanatum, a white rot fungus, was optimized in broth cultures. The fungus was found to have a higher growth rate and higher lectin activity when grown in a medium adjusted to pH 6.5 at 26°C under stationary conditions. Expression of lectin activity started in 5-day-old mycelial culture; maximum activity was expressed after the 15th day of incubation. Among the various carbon and nitrogen sources tested, the carbon source sucrose and the nitrogen source yeast extract support maximum growth and lectin production. Lectin from G. applanatum was purified by ammonium sulfate precipitation and ion exchange chromatography. The purified fraction revealed a single band with a molecular weight of 35.0 kDa. Moreover, carbohydrates such as mannitol, glucose, sucrose, maltose, mannose, galactose, sorbose, and fructose were found to inhibit the hemagglutinating activity of the lectin. The purified lectins from G. applanatum contain cytotoxic and proapoptotic activities against HT-29 colon adenocarcinoma cells.
The aim of this study was to determine the antiulcer activity of Lignosus rhinocerotis in rats. A total of 48 Sprague-Dawley rats were used in ethanol-induced, aspirin-induced, and water immersion-restraint stress-induced ulcer models. Rats were equally divided into 4 groups for each model and orally administered 5 mL/kg distilled water, 20 mg/kg omeprazole, as well as 250 and 500 mg/kg of L. rhinocerotis powder. L. rhinocerotis powder at both 250 and 500 mg/kg doses demonstrated significant (P < 0.05) protection against gastric ulceration in all the induced ulcer models. Histological studies revealed severe damage and hemorrhage of gastric mucosa in the negative control group for all ulcer-induced models. The study suggests that L. rhinocerotis powder possesses dose-dependent antiulcer activity in the gastric mucosa, as ascertained grossly and histologically, compared with the negative control groups.
Rigorous research has been carried out regarding the cultivation of Ganoderma lucidum using different agricultural residues. Nevertheless, large-scale cultivation and the separation of active compounds of G. lucidum are still challenges for local farmers. The objective of this study was to evaluate the use of oil palm waste fibers such as empty fruit bunch fibers and mesocarp fibers as effective substrates for the growth of G. lucidum mycelia to study the possibility of solid-state cultivation and to determine the optimum conditions necessary for the growth of mycelia of this mushroom on these waste fibers. Various parameters such as temperature, pH, humidity, and carbon and nitrogen compositions required for the optimum growth of mycelia have been determined. Oil palm fibers are a vivid source of lignocellulose, and their availability in Malaysia is high compared to that of sawdust. G. lucidum is a wood-rotting fungi that can easily decay and utilize this lignocellulose biomass, a major agricultural waste in Malaysia.
Nonalcoholic fatty liver disease (NAFLD) is currently one of the most common liver diseases worldwide. Lifestyle modifications through the diet are the mainstay of treatment. Auricularia nigricans is a popular edible mushroom known to possess medicinal properties. Gas chromatography-mass spectrometry and liquid chromatography-tandem mass spectrometry analysis indicated that linoleic acid ethyl ester, butyl 9,12-octadecadienoate, 9,12-octadecadienoic acid, ergosta-5,7,22-trien-3-ol, 2(3,4-dihydroxyphenyl)-7-hydroxy-5-benzene propanoic acid, and 3,30-di-0-methyl ellagic acid were present in the A. nigricans ethyl acetate (EA) fraction. The cytotoxicity assay showed that the EA fraction was noncytotoxic to HepG2 cells at concentrations < 100 μg/mL. In the antihepatic steatosis assay, 50 μg/mL of EA fraction caused a decline in absorbance to 0.20 ± 0.02 compared to palmitic acid (PA)-induced cells (0.24 ± 0.02). Furthermore, cells treated with 50 μg/mL and 25 μg/mL of EA fraction contributed an approximately 1.12-fold and 1.08-fold decrease in lipid accumulation compared to PA-induced cells. Coincubation with PA and 25 μg/mL of EA fraction decreased levels of tumor necrosis factor-α, interleukin (IL)-6, IL-8, and monocyte chemoattractant protein-1 to 140.48 ± 8.12, 91.16 ± 2.40, 184.00 ± 22.68, and 935.88 ± 39.36 pg/mL compared to PA-induced cells. The presence of the EA fraction also suppressed the stress-activated protein kinase/Jun amino-terminal kinase, p-38 mitogen-activated protein kinase, nuclear factor-κB, and signal transducer and activator of transcription 3 signaling pathways. In conclusion, these findings suggest that the A. nigricans EA fraction demonstrates antisteatotic effects involving antioxidant capacity, hypolipidemic effects, and anti-inflammatory capacity in the PA-induced NAFLD pathological cell model.
Ophiocordyceps sinensis (=Cordyceps sinensis) has been known for its various medicinal properties, in particular immunomodulatory activities associated with its polysaccharides. In this study, the fruiting body of O. sinensis cultivar OCS02® was investigated for its chemical composition and monosaccharide profile. Cold water extract (CWE) obtained from this fruiting body was fractionated by molecular weight (MW) into high (HMW), medium (MMW), and low (LMW) fractions. Polysaccharides in the extract and fractions were identified as heteroglycans containing mostly glucose and mannose with small amounts of galactose, fucose, arabinose, and xylose. The immunomodulatory potential of these heteroglycans was evaluated by induction of cytokine/chemokine secretion using murine macrophage RAW 264.7. All treatments showed significant modulation of IL-6, IL-9, MIP-2, and TIMP-1, especially for CWE, HMW, and MMW, which might be due to their high ratios of glucose and the presence of protein. Further investigation on the structure-function relationship of these fruiting body polysaccharide fractions is needed to delineate the underlying mechanism of their immunomodulatory effect both in vitro and in vivo.
The edible mushroom Pleurotus giganteus was tested for its effect on adipocyte differentiation and glucose uptake activity in 3T3-L1 cells. The basidiocarps of P. giganteus were soaked in methanol to obtain a crude methanol extract and then fractionated to obtain an ethyl acetate extract. In this study, cell proliferation was measured using an MTT assay, lipid accumulation using an Oil Red O assay, and glucose uptake using a fluorescence glucose uptake assay. Gene expression was measured via real-time polymerase chain reaction analysis with TaqMan primer. Ethyl acetate extract significantly enhanced adipogenic differentiation and glucose uptake in 3T3-L1 adipocytes via the expression of sterol regulatory element-binding protein, peroxisome proliferator-activated receptor γ, and phos-phatidylinositol 3-kinase/Akt. Glucose uptake was facilitated by the highly expressed glucose transporters Glut1 and Glut4. Taken together, these results suggest that P. giganteus ethyl acetate extract has an insulin-sensitizing effect on adipocytes and has potential as an adjuvant for the management of type 2 diabetes.
This study was conducted to evaluate the mycochemical composition and antiglycemic and antioxidant activities of Ganoderma neo-japonicum hot aqueous extracts, prepared at different boiling durations, and polysaccharides isolated from them. Ground basidiocarps of G. neo-japonicum were double-boiled at 100°C for 0.5, 3, or 4 hours, and the antiglycemic activity was assessed by α-amylase and α-glucosidase enzyme inhibition assays. The antioxidant capacity of the crude hot aqueous extracts (AE-1, AE-2, AE-3) was assessed by DPPH and ABTS radical scavenging and ferric-reducing antioxidant power assays. The total phenolics, protein, and sugar in the crude extracts were also determined. The hot aqueous extract (AE-3) containing a significant amount of total sugar and having enhanced antiglycemic and antioxidant activities was selected for polysaccharide isolation. The isolated crude polysaccharide was separated and purified using diethylaminoethyl-cellulose-52 and Sepharose 6B column chromatography. Fourier transform infrared spectroscopy studies of the purified polysaccharide fraction (PF) showed the presence of typical bands corresponding to polysaccharides. The estimated β-glucan concentration in the PF was 39.26%. In general, the PF exhibited significantly lower antioxidant activity than AE-3. Nevertheless, its potency in inhibiting carbohydratehydrolyzing enzymes may have potential in the management of diabetes mellitus.
This study investigated antioxidant and anti-inflammatory properties, and the direct cytotoxic effect of Lignosus rhinocerotis fractions, especially the polysaccharide fraction, on nasopharyngeal carcinoma cells. L. rhinocerotis crude extract was obtained through hot water extraction. The precipitate saturated with 30% ammonium sulfate was purified with ion-exchanged chromatography. Gel permeation chromatography multiangle laser light scattering analysis equipped with light scattering and UV signals revealed two district groups of polymers. A total of four peaks were observed in the total carbohydrate test. Fraction C, which was the second region of the second peak eluted with 0.3 M NaOH, showed the highest integrated molecular weight, whereas fraction E had the lowest integrated molecular weight of 19,790 Da. Fraction A contained the highest β-D-glucan content. Enzymatic analysis showed that most of the polysaccharide fractions contained β-1-3 and β-1-6 skeletal backbones. The peak eluted with 0.6 M NaOH was separated in fraction D (flask 89-92) and fraction E (93-96). The results showed that fraction E expressed higher antioxidant activities than fraction D whereas fraction D expressed higher chelating activity than fraction E. The extract saturated with 30% ammonium sulfate exhibited higher reducing power than the extract saturated with 100% ammonium sulfate. Fractions D and E significantly inhibited the secretion of tumor necrosis factor-α in lipopolysaccharide-stimulated RAW 264.7 macrophages in a dose-dependent manner. There was no apparent difference in the viability of cells exposed or unexposed to L. rhinocerotis fractions.
Oxidative stress (OS) and hypercholesterolemia have been linked with a heightened risk of cardiovascular disease (CVD). Because of the numerous drawbacks of synthetic antioxidants and cholesterol-lowering drugs, natural antioxidative and hypocholesterolemic agents are of immense importance. This study was designed to determine both the OS-attenuating and cholesterol-lowering capacities of the hot water extract (HWE) and of five solvent-solvent-partitioned fractions of Ganoderma lucidum. In vitro antioxidative performance of G. lucidum HWE and fractions was measured through DPPH free radical scavenging, Folin-Ciocalteu assay, lipid peroxidation inhibition, and human low-density lipoprotein (LDL) oxidation inhibition. In vivo antioxidative performance of G. lucidum was assessed by measuring the plasma and liver antioxidative enzymatic activities (catalase, glutathione peroxidase, and superoxide dismutase) in G. lucidum HWE-fed rats. In the CVD tests, the HWE at 200 mg/kg b.w. lowered plasma levels of total cholesterol, triacylglycerol, and LDL cholesterol and increased high-density lipoprotein cholesterol. The current findings indicate the therapeutic potentiality of G. lucidum as an OS-attenuating and hypocholesterolemic agent en route to withstanding CVD complications.
This study investigates the cultivation of Ganoderma lucidum using different agricultural biomasses from Malaysia. Five different combinations of rubber wood sawdust, empty fruit bunch fiber, and mesocarp fiber from oil palm, alone and in combination, were used to cultivate G. lucidum. Although all the substrate combinations worked well to grow the mushroom, the highest biological efficiency was obtained from the combination of empty fruit bunch fiber with sawdust. A total yield of 27% was obtained from empty fruit bunch fiber with sawdust, followed by sawdust (26%), empty fruit bunch fiber (19%), mesocarp fiber with sawdust (19%), and mesocarp fiber (16%). The quality of mushrooms was proved by proximate analysis and detection of phenolic compounds and flavonoids. The antioxidant activity verified by DPPH, ferric-reducing ability of plasma, and ABTS analyses revealed that the empty fruit bunch fiber with sawdust had higher activity than the other substrates.
A central composite design of response surface methodology (RSM) was employed to optimize the extraction time (X1: 266.4-393.6 min) and temperature (X2: 42.9-57.1°C) of Pleurotus ostreatus aqueous extract with high antioxidant activities, namely DPPH radical-scavenging activity, ABTS radical cation inhibition, and ferric reducing/antioxidant power, as well as total phenolic content (TPC). Results showed that the data were adequately fitted into four second-order polynomial models developed by RSM. The extraction time and temperature were found to have significant quadratic effects on antioxidant activities and TPC. The optimal extraction time and temperature were 282.3 min and 42.9°C (DPPH), 393.6 min and 42.9°C (ABTS), 340.4 min and 49.8°C (FRAP), and 347.6 min, 49.7°C (TPC), with corresponding yields of 53.32%, 73.20%, 37.14 mM Fe2+ equivalents/100 g, and 826.33 mg gallic acid equivalents/100 g, respectively. These experimental data were close to their predicted values. The establishment of such a model provides a good experimental basis for employing RSM to optimize the extraction time and temperature for high antioxidant activities from P. ostreatus.
Lignosus rhinocerotis (Cooke) Ryvarden has been reported to possess numerous pharmacological effects. However, little is known about its potential role in mitigating the detrimental effects of oxidative stress. The present study investigated the cytoprotective effects of L. rhinocerotis extracts against hydrogen peroxide (H2O2)-induced oxidative stress of rat pheochromocytoma (PC12) cells. In the pre-treatment model, PC12 cells were pre-treated with aqueous (LRAQ) or ethanolic (LRET) extracts of L. rhinocerotis for 24 h, followed by 30 μM of H2O2 for 24 h. In the co-treatment model, the cells were incubated with LRAQ or LRET and H2O2 for 2 or 24 h to induce oxidative stress. Cell viability, intracellular reactive oxygen species (ROS) levels, mitochondrial membrane potential (MMP), and apoptotic cells with activated caspase-3/7 were quantified. Additionally, LRET was separated into fractions by chromatographic methods prior to analysis by gas chromatography-mass spectrometry (GCMS). 320 μg/ml aqueous extract showed a significant cytoprotective effect of 70.0 ± 22.4% and 133.92 ± 8.8% in the pre-treatment and co-treatment models, respectively, compared to untreated H2O2-challenged cells. LRAQ also showed a reduction (p < 0.05) in the percentage of depolarized cells of 37.6 ± 0.6% at 640 ug/ml and 53.4 ± 4.5% at 320 ug/ml in the pre-treatment and co-treatment models, respectively, compared to untreated H2O2-challenged cells. LRAQ or LRET showed a reduction (p < 0.01) in caspase 3/7 activity compared to untreated H2O2-challenged cells in the co-treatment model. However, LRAQ or LRET did not reduce excessive ROS formation (p > 0.05). The cytoprotective effects could be attributed to the presence of fatty acids, phenols, phytosterols, and dicarboxylic acids. In conclusion, L. rhinocerotis extracts demonstrated cytoprotective effects against H2O2-induced oxidative stress in an in vitro model, contributing to the maintenance of cellular integrity through the regulation of mitochondrial function and apoptosis.
Black jelly mushroom (Auricularia polytricha) is a well-known Chinese traditional food that has therapeutic effects. This study evaluated the effects of different cooking methods (boiling, steaming, microwaving, and stir-frying) on the physicochemical characteristics (i.e., total phenolic content, antioxidants, α,α-diphenyl-β-picryl-hydrazyl [DPPH] free radical scavenging, and ferric reducing antioxidant power [FRAP]) along with color, texture, moisture, and sensory properties of black jelly mushrooms. Lightness (L*) was significantly lower for the stir-frying method (29.93) compared to the control (34.62). Stir-fried mushrooms had significantly lower firmness force (texture) and moisture content (80.13 N and 61.98%, respectively) compared to the control (2000.37 N and 86.52%). The steaming method contributed significantly higher total phenolic content (11.23 mg gallic acid equivalents/g) and antioxidant activity measured using the FRAP (33.54 mg Trolox equivalents/g) and DPPH (90.41% inhibition) assays compared to the respective controls.
The present study aims to assess the antioxidant activities (AOA) and total phenolic content (TPC) of water extracts of selected edible wild mushrooms: Pleurotus porrigens, Schizophyllum commune, Hygrocybe conica, and Lentinus ciliatus. The AOA were evaluated against DPPH radical and ABTS radical cation scavenging ability, ferric-reducing antioxidant power (FRAP) and beta-carotene-linoleate bleaching (beta-CB) assays, and the Folin-Ciocalteu method for TPC. BHA was used as reference. P. porrigens showed significantly higher (p < 0.05) DPPH* scavenging ability (90.78 +/- 0.30%) and FRAP (6.37 +/- 0.22 mM FE/100g), while Sch. commune showed significantly higher (p < 0.05) ABTS*+ inhibition activity (94.96 +/- 0.70%) and beta-CB inhibition activity (94.18 +/- 0.17%), respectively. TPC was found in a descending order of P. poriggens > L. ciliatus = Pleurotus ostreatus (cultivated) > H. conica = Sch. commune. Positive correlation was observed between the AOA and TPC. When compared to BHA (2 mM), P. porrigens showed significantly higher (p < 0.05) DPPH* scavenging ability and reducing power, while Sch. commune showed comparable DPPH* scavenging ability and ABTS*+ inhibition activity. All the mushrooms have better ABTS*+ inhibition activity than BHA (1 mM). The beta-CB inhibition activity of BHA was significantly higher than those of edible wild mushrooms. The water extracts of edible wild mushrooms showed potent antioxidant activities compared to BHA to a certain extent.