Optimization of Antioxidant Activities and Intracellular Polysaccharide Contents Using Agaricus bisporus Extract as Elicitor in Submerged Fermenting Ganoderma lucidum

Original strains of Ganoderma lucidum ( MZKI G97) and Grifola frondosa (GF3) isolated from Slovenian forests were cultivated using submerged and solid state cultivation. In 14 days submerged Ganoderma lucidum fed batch cultivation extracellular (1,7 g/L) and intracellular (0.45 g/L) polysaccharide fractions were isolated, up to 17.0 g/L dry fungal biomass was produced, while in 28 days Grifola frondosa fed batch cultivation 3.65 g/L of extracellular and 1.30 g/L intracellular polysaccharide and 15.2 g/L dry biomass was produced. In Ganoderma lucidum solid state cultivation in 18 days 5.77 mg /g extracellular and 1.45 mg /g intracellular polysaccharide was produced while in Grifola frondosa in 38 days cultivation of 0.70 extracellular and 3.80 mg/g of intracellular polysaccharide were produced. The isolated polysaccharides were mainly -D-glucanes. Immunostimulatory effects of isolates were tested on induction of cytokine (TNF- , IFN- and IL12) synthesis in primary cultures of human mononuclear cells (PBMC) isolated from a buffy coat.

Mushrooms, renowned for their nutritional value and bioactive compounds, offer potential health benefits, including antioxidants and anti-aging properties. Aging, characterized by cellular and tissue decline, is often associated with autophagy dysfunction, a crucial cellular cleaning process. This study aimed to investigate the neuroprotective, hepatoprotective, and antimicrobial properties of extracts from four medicinal and edible mushrooms: Ganoderma lucidum, Hericium erinaceus, Pleurotus ostreatus, and Agaricus bisporus. The protein, total phenol, and flavonoid content of mushroom extracts were determined. Aging was induced with 120 mg/kg D-galactose and treated with 500 mg/kg mushroom extracts. The study evaluated liver enzyme levels, histopathological changes in liver and brain tissues, gene expression correlated to neurodegeneration (SEPT5-SV2B-ATXN2-PARK2), telomere length, and immunomodulatory and pro-inflammatory (IL-2-IL-4-IL-6) gene expression pathways. Additionally, the antimicrobial potential of mushroom extracts was assessed against several bacteria (Lysinibacillus odyssey, Lysinibacillus fusiformis, Klebsiella oxytoca, and Escherichia coli) using agar well diffusion and lowest minimum inhibitory concentration (MIC) methods. By exploring these diverse aspects, this study aimed to provide a foundation for a better understanding of the potential of mushrooms as natural neuroprotective, hepatoprotective, and antimicrobial agents and their potential applications in human health. Results indicated that all mushroom extracts effectively mitigated oxidative stress. Agaricus bisporus exhibited the highest protein and flavonoid content, and Pleurotus ostreatus displayed the highest phenolic content. Notably, Hericium erinaceus and Ganoderma lucidum extracts demonstrated significant neuroprotective and hepatoprotective properties against D-galactose-induced aging, as evidenced by histopathological examination. All extracts exhibited a significant decrease (p < 0.001) in liver function (serum levels of aspartate aminotransferase (GOT) and alanine aminotransferase (GPT)) and showed immunomodulatory and anti-inflammatory effects, characterized by upregulated IL-2 and IL-4 gene expression and downregulated IL-6 gene expression. Hericium erinaceus demonstrated the most pronounced upregulation (p < 0.001) of SEPT5, SV2B, and telomere length gene expression, suggesting potential anti-aging effects. Furthermore, all mushroom extracts displayed antimicrobial activity against the tested bacterial strains, except Hericium erinaceus, which exhibited antibacterial activity solely against E. coli. Agaricus bisporus exhibited the largest inhibition zones (22 ± 0.06 mm) against Lysinibacillus odyssey, while Hericium erinaceus displayed the largest inhibition zone against E. coli. The MIC value was observed with Agaricus bisporus extract against Lysinibacillus odyssey (1.95 ± 0.16 mg/mL). Lysinibacillus fusiformis exhibited the highest resistance to the tested mushroom extracts. These findings suggest that these edible and medicinal mushrooms possess a wide range of health-promoting properties, including neuroprotective, hepatoprotective, and antimicrobial activities. Further research is needed to fully understand the underlying mechanisms and optimize applications. However, our results provide a strong foundation for exploring these mushrooms as potential natural agents that promote overall health and combat age-related decline.

Oxidative stress can disrupt the body's ability to fight harmful free radicals, leading to premature aging and various health complications. This study investigated the antioxidant and anti-aging properties of four medicinal and edible mushrooms: Ganoderma lucidum, Hericium erinaceus, Pleurotus ostreatus, and Agaricus bisporus. The antioxidant activity of mushroom extracts was evaluated using (DPPH-ABTS-Reducing power). The anti-aging effects were assessed using Human Skin Fibroblasts (HSF) cells subjected to D-galactose-induced aging (30 g/L/72 h) and treated with mushroom extracts (0.03-0.25 mg/mL/72 h). The results demonstrated that all mushrooms have significant antioxidant and anti-aging properties, with low concentrations of extracts (0.03 mg/mL) effectively promoting cell proliferation at an 87% rate in the Agaricus bisporus extract, enhancing cell cycle progression by reducing the arrested cells in the G0/G1 phase to 75%, and promoting DNA synthesis in S phase by more than 16.36% in the Hericium erinaceus extract. Additionally, the extracts reduced DNA damage and Reactive Oxygen Species (ROS) levels, protecting cells from oxidative stress and potentially contributing to anti-aging effects. The mushrooms also exhibited immunomodulatory and anti-inflammatory effects by upregulating the IL-2, IL-4, and downregulating IL-6 expression, indicating their potential to promote general health. These findings suggest the potential of mushroom extracts as natural agents for reducing the negative effects of aging while promoting cellular health. Further research is required to explore the specific bioactive compounds responsible for these beneficial effects and to evaluate their efficacy in vivo.

In the present work crude Agaricus bisporus extract (ABE) has been prepared and characterized by its tyrosinase activity, protein composition and substrate specificity. The presence of mushroom tyrosinase (PPO3) in ABE has been confirmed using two-dimensional electrophoresis, followed by MALDI TOF/TOF MS-based analysis. GH27 alpha-glucosidases, GH47 alpha-mannosidases, GH20 hexosaminidases, and alkaline phosphatases have been also detected in ABE. ABE substrate specificity has been studied using 19 phenolic compounds: polyphenols (catechol, gallic, caffeic, chlorogenic, and ferulic acids, quercetin, rutin, dihydroquercetin, l-dihydroxyphenylalanine, resorcinol, propyl gallate) and monophenols (l-tyrosine, phenol, p-nitrophenol, o-nitrophenol, guaiacol, o-cresol, m-cresol, p-cresol). The comparison of ABE substrate specificity and affinity to the corresponding parameters of purified A. bisporus tyrosinase has revealed no major differences. The conditions for spectrophotometric determination have been chosen and the analytical procedures for determination of 1.4 × 10−4–1.0 × 10−3 M l-tyrosine, 3.1 × 10−6–1.0 × 10−4 M phenol, 5.4 × 10−5–1.0 × 10−3 M catechol, 8.5 × 10−5–1.0 × 10−3 M caffeic acid, 1.5 × 10−4–7.5 × 10−4 M chlorogenic acid, 6.8 × 10−5–1.0 × 10−3 M l-DOPA have been proposed. The procedures have been applied for the determination of l-tyrosine in food supplements, l-DOPA in synthetic serum, and phenol in waste water from the food manufacturing plant. Thus, we have demonstrated the possibility of using ABE as a substitute for tyrosinase in such analytical applications, as food supplements, medical and environmental analysis.

This study was designed to assess the nephroprotective effects of Pleurotus ostreatus and Agaricus bisporus aqueous extracts and carvedilol on hyperoxaluria-induced urolithiasis and to scrutinize the possible roles of NF-κB, p53, Bcl-2, Bax and Bak. Phytochemical screening and GC-MS analysis of mushrooms’ aqueous extracts were also performed and revealed the presence of multiple antioxidant and anti-inflammatory components. Hyperoxaluria was induced in Wistar rats through the addition of 0.75% (v/v) ethylene glycol in drinking water for nine weeks. The ethylene glycol-administered rats were orally treated with Pleurotus ostreatus and Agaricus bisporus aqueous extracts (100 mg/kg) and carvedilol (30 mg/kg) daily during the last seven weeks. The study showed that Pleurotus ostreatus, Agaricus bisporus and carvedilol all successfully inhibited ethylene glycol-induced histological perturbations and the elevation of serum creatinine, serum urea, serum and urinary uric acid, serum, urinary and kidney oxalate, urine specific gravity, kidney calcium, kidney NF-κB, NF-κB p65, NF-κB p50, p53, Bax and Bak expressions as well as serum TNF-α and IL-1β levels. Moreover, the treatment decreased the reduction in urinary creatinine, urinary urea, ratios of urinary creatinine to serum creatinine and urinary urea to serum urea, Fex Urea and Bcl-2 expression in kidney. In conclusion, although Pleurotus ostreatus and Agaricus bisporus extracts and carvedilol all significantly inhibited the progression of nephrolithiasis and showed nephroprotective effects against ethylene glycol-induced kidney dysfunction, Pleurotus ostreatus and Agaricus bisporus seemed to be more effective than carvedilol. Moreover, the nephroprotective effects may be mediated via affecting NF-κB activation, extrinsic apoptosis and intrinsic apoptosis pathways.

Wild-cultivated medicinal mushroom Ganoderma lucidum was morphologically identified and sequenced using phylogenetic software. In submerged-liquid fermentation (SLF), biomass, exopolysaccharide (EPS) and intracellular polysaccharide (IPS) production of the identified G. lucidum was optimised based on initial pH, starting glucose concentration and agitation rate parameters using response surface methodology (RSM). Molecularly, the G. lucidum strain QRS 5120 generated 637 base pairs, which was commensurate with related Ganoderma species. In RSM, by applying central composite design (CCD), a polynomial model was fitted to the experimental data and was found to be significant in all parameters investigated. The strongest effect (p < 0.0001) was observed for initial pH for biomass, EPS and IPS production, while agitation showed a significant value (p < 0.005) for biomass. By applying the optimized conditions, the model was validated and generated 5.12 g/L of biomass (initial pH 4.01, 32.09 g/L of glucose and 102 rpm), 2.49 g/L EPS (initial pH 4, 24.25 g/L of glucose and 110 rpm) and 1.52 g/L of IPS (and initial pH 4, 40.43 g/L of glucose, 103 rpm) in 500 mL shake flask fermentation. The optimized parameters can be upscaled for efficient biomass, EPS and IPS production using G. lucidum.

Article history:Received 16 July 2014Accepted 20 February 2015Available online 1 April 2015Keywords:β-GlucanAcute oral toxicityMacrophage cell linePhagocytosisStirred tank reactorTotal carbohydrate Background: The role of polysaccharides isolated from the Ganoderma species of fungi in innate immunity hasrecently become a topic of research. Although some work has been conducted concerning Ganoderma lucidum,the characteristics of polysaccharides isolated from Ganoderma neojaponicum (Imazeki) as immunomodulatoryagents are largely unknown. The aims for this study were to isolate and characterize the intracellularpolysaccharides (IPSs) and extracellular polysaccharides (EPSs) of G. neojaponicum from STR reactor.Results:TheproductionofEPSandIPSwasoptimizedonday4ofthecultivationtimein2LSTRreactorbasedontheamountof biomass yield,total carbohydrate, β-glucan and α-glucancontent. Further analysis, both theEPSsand IPSs showed the enhancement on proliferation and increment of phagocytosis activities of macrophage(RAW264.7) cell lines. Using an oral toxicity test, we also observed that 2000 mg/kg body weight/day dosageof dried G. neojaponicum mycelium does not cause any significant toxic effects on Sprague–Dawley rats in 14 dof administration.Conclusion: The findingsof this study indicate that the IPSs and EPSs of G. neojaponicum have the potential to beused as immunomodulating agents to stimulate the innate immune system for fighting infectious diseases. Thepolysaccharides from G. neojaponicum have to be further commercially explored as an alternative formedicinal Ganoderma variety of G. lucidum production.©2015Pontificia Universidad Catolica de Valparaiso. Production and hosting by Elsevier B.V. All rights reserved.

The objective of this study was to increase the intracellular polysaccharide yield of Ganoderma lucidum. The accordingly optimized fermentation medium by central composite design method contains glucose 40gL-1, yeast powder 12gL-1, potassium dihydrogen phosphate 3gL-1, initial pH5.5, and inoculum size 10mL100mL-1. Under this condition, the predicted value of intracellular polysaccharide yield was 2.03gL-1. Shake flask experiments confirmed that the average intracellular polysaccharide yield was 1.98gL-1 similar to the predicted value. The yields of intracellular polysaccharides in the 5-L and 50-L fermentors were 2.59gL-1 and 2.65gL-1, respectively. The molecular weight distribution of intracellular and extracellular polysaccharides obtained was determined by HPSEC-MALLS-RI. The results showed that the weight-average molecular weight of component 1 in the intracellular crude polysaccharide was 4.695 × 106Da and the mass fraction was 58%. The weight-average molecular weight of component 2 in the extracellular polysaccharide was 5.554 × 104Da. The mass fraction was 94.9%. The liquid submerged fermentation process of G. lucidum mycelium obtained from this study has effectively increased the yield of intracellular polysaccharides. Its intracellular and extracellular polysaccharides have good immunological activity. Conceivably, the optimized process can be applied for the large-scale production.

A research had been done on antibacterial activity of extract and the aim was to determine the antibacterial activity and inhibitory potency. The research was started by screening test using Bacillus subtilis, Escherichia coli, Pseudomonas aeruginosa, Salmonella typhi, Staphylococcus epidermidis, Shigella dysenteriae, Staphylococcus aureus, Streptococcus mutans and Vibrio cholerae. Results showed that the Agaricus bisporus ekstract inhibited Bacillus subtilis, Shigella dysenteriae, Staphylococcus aureus, and Vibrio cholera at concentrations 0,1% and 1%. The results obtained Minimum Inhibitory Concentration (MIC) Agaricus bisporus extract against at concentrations of 0,1%. In the Minimum Bactericidal Concentration (MBC) test Agaricus bisporus extract against at concentrations of 0,8%. Agaricus bisporus extract has antibacterial activity wich was indicated by the inhibitory zone diameter. The biggest inhibitory zone diameter of Agaricus bisporus contained in bacteria Bacillus subtilis on the concentration of 6.4% (13 mm)

In this study, one strain of Beauveria caledonica was isolated from wild fruiting bodies collected from Guizhou Province, China, and its species identification, biological characteristics, domestication, and cultivation methods were studied along with polysaccharide and adenosine content analysis. The mycelia were identified by ITS sequencing, and the fruiting bodies of B. caledonica were domestically cultivated for the first time using wheat and rice as basic cultivation media. The carbon sources, nitrogen sources, cultivation temperatures, and pH for mycelial growth were optimized through single-factor experiments and response surface methodology (RSM) experiments. The polysaccharide content was detected by the phenol-sulfuric acid method, and the adenosine content was measured by high-performance liquid chromatography (HPLC). The results confirmed that the identified mycelia were B. caledonica. The optimum medium for solid culture was 25.8 g/L glycerol, 10.9 g/L yeast extract, 1 g/L MgSO4·7H2O, 1 g/L KH2PO4, 10 mg/L vitamin B1, and 20 g/L agar; the optimum pH was 6.5, and the optimum culture temperature was 25 °C. The optimal liquid culture medium was 26.2 g/L glycerol, 11.1 g/L yeast extract, 1 g/L MgSO4·7H2O, 1 g/L KH2PO4, and 10 mg/L vitamin B1; the mycelia grew well at pH 6.6 and 25 °C. The average biological efficiencies of fruiting bodies on wheat and rice as culture media were 1.880% and 2.115%, respectively; the polysaccharide contents of fruiting bodies on the two media were 6.635% and 9.264%, respectively, while the adenosine contents were 0.145% and 0.150%, respectively. This study provides a valuable reference for further artificial cultivation and utilization of B. caledonica by investigating its biological characteristics, cultivation conditions for artificial domestication, and polysaccharide and adenosine contents in cultivated fruiting bodies.

Basidiomycetes, known for their production of bioactive compounds, traditionally use simple sugars for fermentation. However, their ability to degrade complex plant polysaccharides through enzyme secretion presents potential for the use of renewable raw materials. This study focused on the optimization of exopolysaccharide (EPS) production and efficient substrate consumption by Ganoderma lucidum using response surface methodology (RSM). Using an optimized medium containing 15 g⋅l-1 wheat starch, 0.375 g⋅l-1 NH4Cl, and 0.75 g⋅l-1 CaCl2 (C/N ratio of 40), a significant increase in EPS concentration from 121.1±10.2mg⋅l-1 to 229.0±20.3mg⋅l-1 and starch degradation degree (SDD) from 9.1% to 57.6% was achieved after 9 d of submerged cultivation. Scale-up experiments were conducted in both column and stirred tank bioreactors, employing submerged and immobilized cultivation modes. Submerged cultivation in the column bioreactor yielded the highest process desirability of 0.56, achieving EPS concentration of 192.5±5.4mg⋅l-1 and 60.2% SDD within 7 d. These results highlight the potential of the used column bioreactor for efficient and rapid EPS production. Notably, bioreactor experiments revealed local maxima in EPS content at specific time points, suggesting that cell wall degradation, potentially induced by shear stress, may contribute to the release of polysaccharides into the culture broth.

Ganoderma lucidum (GL) is a well-known medicinal mushroom that has been extensively cultivated. Our previous study has shown that abundant Trichoderma colonies grow on the casing soil surface, posing cultivation obstacles for GL. However, an understanding of species-level characteristics of Trichoderma strains and their adverse effects on GL growth is limited. This study aimed to investigate the diversity and potential effects of Trichoderma from GL-cultivated soils. Over 700 Trichoderma isolates were collected from two trails in Longquan Country, southeast China. Eight Trichoderma species, including T. atrioviride, T. guizhouense, T. hamatum, T. harzianum, T. koningiopsis, T. pleuroticola, T. sp. irale, and T. virens, were identified based on the combination alignment of tef-1α and rpb2 sequences. The number of Trichoderma colonies increased dramatically during GL cultivation, with an increase of 9.2-fold in the Lanju trail. T. virens accounted for the most colonies (33.33 and 32.50% in Lanju and Chengbei, respectively) at the end of GL cultivation. The Trichoderma species growth varied but was satisfactory under different temperature or pH conditions. Moreover, Trichoderma species showed different adverse effects on GL growth. The non-volatile metabolites from T. virens and volatile metabolites from T. atroviride displayed the strongest antagonistic activity. Furthermore, the volatile 6-pentyl-2H-pyran-2-one (6-PP) showed a significant inhibitory effect on GL growth with an 8.79 μl mL-1 headspace of 50% effective concentration. The different Trichoderma spp. produced different amounts of 6-PP. The most efficient 6-PP producer was T. atroviride. To the best of our knowledge, this study is the first to demonstrate the abundance of competitive Trichoderma species associated with GL cultivation. Our results would contribute to.

Ganoderma lucidum es un hongo macromiceto reconocido por sus propiedades medicinales y su contenido de compuestos bioactivos que incluyen polisacáridos, triterpenoides, proteínas inmunomoduladoras, entre otros, lo que ha generado un incremento notable en su producción. La mayoría de especies de hongos responden y se adaptan a diversas señales ambientales incluida la luz, que favorece su productividad, tanto en calidad como en cantidad al estar estrechamente relacionada con la formación de cuerpos fructíferos. Por tal razón, el objetivo de este estudio fue evaluar la eficiencia biológica (EB) y la tasa de producción (TP) como parámetros de productividad del cultivo sólido de Ganoderma lucidum bajo irradiación de los sustratos con luz emitida por diodos azules (LED) con dos periodos de foto-estímulo de 12 y 24 h durante todas las fases de cultivo para inducir el crecimiento micelial y la formación de los cuerpos fructíferos. Se aplicaron parámetros convencionales para el crecimiento y desarrollo del hongo en las etapas de producción. Para la formulación de los sustratos, se emplearon residuos agroindustriales y materiales lignocelulósicos. El diámetro de los cuerpos fructíferos sometidos a tratamientos con luz azul fue mayor que los exhibidos a luz blanca fluorescente (Testigo). Los resultados muestran que el cultivo de Ganoderma lucidum con exposición a la luz azul es útil para la inducción de cuerpos fructíferos de alta calidad, logrando una disminución del periodo de fermentación en 16 días para el foto-estímulo de 24 h con EB de 28,04% y TP de 0,64.

This research was conducted to examine Agaricus bisporus and Auricularia auricula crude extract using different solvents (water, ethanol, and methanol) on infrared spectroscopy absorbance during extraction and the impact on broiler carcass. Agaricus bisporus and Auricularia auricula crude extracts were scanned using fourier transform infrared (FT-IR) spectroscopy. Each mushroom crude extract was chosen and applied into broiler diets as feed additive at 0%, 0.4%, 0.8%, 1.2%, and compared zinc bacitracin inclusion. Variable measured were final live weight, carcass yield, breast meat yield, and abdominal fat yield of broiler. Two hundred and forty day-old chicks were randomly allocated into eight dietary treatments, each treatment was replicated three times with ten chicks for each pen. Diets and water were offered ad libitum. Methanolic extract showed monosaccharide absorption peak in fingerprint region at wavelength 890 cm-1, 930 cm-1, 1050 cm-1, 1150 cm-1 which indicates alpha and beta linkage than the others solvent. Even so, dietary inclusion of methanolic extracts of Agaricus bisporus and Auricularia auricula did not show any effect on final live weight and the yiled of carcass, breast meat and abdominal fat of broiler. In conclusion, methanolic extraction is effective to extract monosaccharides with α- and β- linkages from Agaricus bisporus and Auricularia auricula, while the dietary inclusion of methanolic extracts of both edible mushroom and zinc bacitracin has no effect on carcass quality of broilerin broiler diets did not show differences between treatments as well as zinc bacitracin group.

Branched polysaccharides have been isolated from basidiomycete raw materials of locally growing and cultivated Ganoderma lucidum. It has been found that the isolated fractions contain branched polysaccharides in the form of complexes with melanin. After purification of polysaccharides by ion exchange chromatography from locally growing and cultivated basidial raw materials, two fractions have been obtained: neutral polysaccharides of locally growing Ganoderma lucidum (GW-1), cultivated Ganoderma lucidum (GWL-1) with a yield of 25.71 and 29.85%, respectively, and anionic polysaccharides of locally growing Ganoderma lucidum (GW-2), cultivated Ganoderma lucidum (GWL-2), with a yield of 5.26 and 4.19%. The physicochemical properties of the obtained samples have been studied by IR and UV spectroscopies. The purity degree of fractions of branched polysaccharides has been determined. Using gas chromatography, one-dimensional (13C NMR, 1H NMR), and two-dimensional (COSY, TOCSY, HSQC, HMBC, NOESY) NMR spectroscopies, the compositions and molecular structures of the obtained polysaccharide samples have been determined. The results showed that the isolated and purified polysaccharides are β-glucan-type branched polysaccharides that have branch point (1,4,6)- and (1,3,6)-linked glucopyranose residues.