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Endoglucanases from glycoside hydrolase family 5 (GH5) are the key enzymes in degradation of diverse plant polysaccharides. Present study reports purification, characterization and partial sequencing of novel thermostable GH5 family endoglucanase from a newly isolated brown rot fungi Fomitopsis meliae CFA 2. Endoglucanase was purified 34.18 fold with a specific activity of 302.90 U/mg. The molecular weight of the endoglucanase was 37.87 kDa as determined by SDS PAGE. LC MS/MS analysis identified the protein to be a member of GH5_5 family. The temperature and pH optima for endoglucanase activity were 70 °C and 4.8, respectively. The enzyme catalyzed the hydrolysis of carboxymethyl-cellulose with a Km of 12.0 mg/ml, Vmax of 556.58 μmol/min/mg and Kcat of 129.41/sec. The enzyme was stimulated by Zn+2 and K+ metal ions and DTT. Half-life (t1/2) for endoglucanase was found to be 11.36 h with decimal reduction time (D) of 37.75 h at 70 °C. The activation energy for endoglucanase was found to be 30.76 kJ/mol (50 °C–70 °C). Looking at the results, the endoglucanase from Fomitopsis meliae CFA 2 seems to be a promising thermostable enzyme which may be applicable in applications like biomass hydrolysis.

To efficiently decompose biomass, fungi have developed various enzymatic and non-enzymatic strategies and are a source of versatile biocatalysts. The endoglucanases in glycosyl hydrolase CAZy family 45 (GH45) are known for their small size, a high thermostability and a broad substrate specificity that has been employed in textile and detergent industries. Here we report the heterologous expression and characterisation of an GH45 endoglucanase from the brown rot Fomitopsis pinicola and its direct comparison to an already characterised GH45 from the white rot Phanerochaete chrysosporium. Both enzymes were recombinantly expressed in Pichia pastoris and purified by two chromatographic steps. The biochemical characterisation highlighted the acidophilic character, with an optimal pH of 4, and a preference for amorphous substrates as carboxymethyl cellulose (CMC) and substrates containing β-1,4-glucans rather than the previously reported β-1,3/1,4-glucans lichenan and β-glucan. The dominating products from β-1,4-glucans were C3-C6 oligosaccharides, whereas from β-1,3/1,4-glucans glucose was the main reaction product. From the characterisation no differences between the brown rot and the white rot GH45 was evident.

Ceriporiopsis subvermispora (C. subvermispora) is a selective degrader of lignin in the woody biomass. Glutathione S-transferases (GSTs) are multifunctional enzymes that play important roles in cellular detoxification and metabolism. The crystal structures of a GST of C. subvermispora, CsGST83044, in GSH-free and -bound forms were solved at 1.95 and 2.19 A resolution, respectively. The structure of the GSH-bound form revealed that CsGST83044 can be categorized as an atypical-type of GST. In the GSH-bound form of CsGST83044, Asn22, Asn24, and Tyr46 are located closest to the sulfur atom and form hydrogen bonds with the thiol group. The functional mutagenesis indicated that they are critical for the enzymatic activities of CsGST83044. The critical residues of an atypical-type GST belonging to the GSTFuA class were revealed for the first time. A previous study indicated that CsGST83044 and another GST, CsGST63524, differ in substrate preference; CsGST83044 prefers smaller substrates than CsGST63524 for its esterase activity. The GSH-bound pocket of CsGST83044 turns out to be small, which may explain the preference for smaller substrates. Protein engineering of GSTs of C. subvermispora in the light of the obtained insight may pave a path in the future for utilization of the woody biomass.

Brown-rot fungi are the wood-decay basidiomycetes and have ability to break down plant cell wall carbohydrates. It has been suggested that degradation of pectin is important for the initial stages of brown rot. We purified an endo-polygalacturonase (FpPG28A) from the brown-rot fungus Fomitopsis palustris, analysis of the predicted amino acid sequence indicated that FpPG28A belongs to GH family 28. The highest activity of purified FpPG28A was observed at 60 °C in 50 mM sodium acetate buffer (pH 5.0); this activity was highly specific for polygalacturonic acid chains. However, calcium polygalacturonate gel was not degraded by FpPG28A under those optimal conditions. We observed that calcium polygalacturonate gel was readily degraded by the enzyme in the oxalate buffer. Furthermore, the thermostability of FpPG28A was elevated in oxalate buffer at pH 3.0. These results indicated that oxalate has an important role in the degradation of woody pectin by FpPG28A.

Laccase is predominantly found in lignin degrading filamentous white rot fungi, where it is involved in the oxidative degradation of this recalcitrant heteropolymer. In brown rot fungi it is much less prevalent: laccases from only a few brown rots have been detected and only two have been characterized. This study tries to understand the role of this ligninolytic enzyme in brown rots by investigating the catalytic properties of laccases secreted by Fomitopsis pinicola FP58527 SS1. When grown on either poplar or spruce wood blocks, several laccases were detected in the secretome. Two of them (FpLcc1 and FpLcc2) were heterologously produced using Trichoderma reesei QM9414 Δxyr1 as expression host and purified to homogeneity by consecutive steps of hydrophobic interaction, anion exchange and size exclusion chromatography. With the substrates 2,2-azino-bis(3-ethylthiazoline-6-sulfonate) (ABTS), 2,6-dimethoxyphenol (2,6-DMP) and guaiacol both laccases showed similar, low pH-optima below 3 for ABTS and 2,6-DMP and at pH 3.5 for guaiacol which is at the acidic end of laccases isolated from white rot fungi. The determined KM values were low while kcat values measured at acidic conditions were comparable to those reported for other laccases from white rot fungi. While both enzymes showed a moderate decrease in activity in the presence of oxalic and citric acid FpLcc2 was activated by acetic acid up to 3.7 times. This activation effect is much more pronounced at pH 5.0 compared to pH 3.0 and could already be observed at a concentration of 1 mM acetic acid.

Twelve undescribed lanostane-type triterpenes, and twenty-two known triterpenes were isolated and identified from a medicinal bracket fungus Fomitopsis pinicola (Sw.) P. Karst. The structures of these compounds were determined by spectroscopic and spectrometric analyses. The antiinflammatory potential of thirty-two triterpene compounds was evaluated using neutrophils as an assay model, and pinicolasin J was the most potent inhibitor of superoxide anion generation and elastase release, with IC50 values of 1.81 ± 0.44 and 2.50 ± 0.64 μM, respectively. This study provides scientific insight into the nutritional supplement value and medicinal development of Fomitopsis pinicola.

Significant concentrations of pharmaceuticals and personal care products (PPCPs) have been detected in aquatic environment. Fungal enzymatic processes can oxidize these persistent PPCPs; thus, these processes have attracted considerable attention from the scientific community. Here, we evaluated the efficacy of the removal of PPCPs using native fungal enzymes derived from Bjerkandera spp. TBB-03 under various conditions. Among the eight lignocellulosic substrates, ash, which showed the highest laccase production, was selected as the sole enzyme inducer. TBB-03 laccase was found to exhibit remarkable stability under varied pH and temperature conditions. Acetaminophen and bisphenol A were effectively removed by TBB-03 laccase under various conditions, except at pH 8. Although TBB-03 laccase could not efficiently remove single-state sulfamethoxazole directly, a 22% of improvement in sulfamethoxazole removal was observed in the presence of acetaminophen. Overall, our proposed approach showed that Bjerkandera adusta TBB-03 can be potentially applied for further research regarding PPCP remediation.

Fungal mycelia can eliminate almost all cocultured cyanobacterial cells within a short time. However, molecular mechanisms of algicidal fungi are poorly understood. In this study, a time‐course transcriptomic analysis of algicidal fungus Bjerkandera adusta T1 was applied to investigate gene expression and regulation. A total of 132, 300, 422, and 823 differentially expressed genes (DEGs) were identified at 6, 12, 24, and 48 hr, respectively. Most DEGs exhibited high endopeptidase activity, cellulose catabolic process, and transmembrane transporter activity by using Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses. Many decomposition genes encoding endopeptidases were induced a little later in B. adusta T1 when compared with previously investigated algicidal fungus Trametes versicolor F21a. Besides, the accumulated expression of Polysaccharide lyases8 (PL8) gene with peptidoglycan and alginate decomposition abilities was greatly delayed in B. adusta T1 relative to T. versicolor F21a. It was implied that endopeptidases and enzymes of PL8 might be responsible for the strong algicidal ability of B. adusta T1 as well as T. versicolor F21a., Algicidal process of fungus Bjerkandera adusta T1 was investigated by a time‐course transcriptomic analysis. The results showed that most differentially expressed genes exhibited high endopeptidase activity, cellulose catabolic process, and transmembrane transporter activity. Comparison of gene expression between B. adusta T1 and Trametes versicolor F21a during the algicidal process suggested that endopeptidases together with enzymes of the polysaccharide lyases family 8 might be responsible for the strong algicidal activity of B. adusta T1 as well as T. versicolor F21a.

Fermentation is a metabolic process that converts sugars into acids, gases, or alcohol. This process occurs in yeasts and bacteria, as well as in muscle cells when faced with a lack of oxygen. In this paper, isolation, culture, purification and extracellular polysaccharides of strain Fomes fomentarius were studied. Extraction of polysaccharides from a culture based on F. fomentarius extracellular polysaccharides, extracellular polysaccharides fermentation experiments was optimized and compared, the optimal fermentation method was obtained; extracellular polysaccharides were sulfated, phosphorylated experiments, selenium acidified, discussed the preparation of derivative polysaccharides and microscopic detection, and finally studied extracellular polysaccharides on DPPH, The scavenging ability, superoxide anion radical and hydroxyl radical scavenging ability of the derived polysaccharides were compared. The results showed that the extracellular polysaccharide and derivatized polysaccharide of F. fomentarius had certain antioxidant activity.

Medicinal mushrooms of the order Polyporales have a long history of use, which is evidenced by the finding of dissected fruiting bodies with Ötzi, who lived over 5000 years ago. Because of its valuable biological properties and its use in 18th and 19th-century pharmacy, Fomitopsis officinalis used to be mass-collected. Moreover, the large demand for larch wood and non-wood materials (resin) caused an excessive exploitation of larch forests, which directly contributed to the disappearance of F. officinalis from its natural environment. The qualities of medicinal preparations obtained from the F. officinalis fruiting bodies are determined by the unique composition of its bioactive compounds, such as: triterpenoids, polysaccharides, organic acids, coumarins and phenolic compounds. It has been proved that both crude extracts and the compounds isolated from F. officinalis have a wide spectrum of therapeutic effects, including anti-inflammatory, cytotoxic, and antimicrobial effects.

A main cellular functional module that becomes dysfunctional during aging is the proteostasis network. In the present study, we show that benzoic acid derivatives isolated from Bjerkandera adusta promote the activity of the two main protein degradation systems, namely the ubiquitin-proteasome (UPP) and especially the autophagy-lysosome pathway (ALP) in human foreskin fibroblasts. Our findings were further supported by in silico studies, where all compounds were found to be putative binders of both cathepsins B and L. Among them, compound 3 (3-chloro-4-methoxybenzoic acid) showed the most potent interaction with both enzymes, which justifies the strong activation of cathepsins B and L (467.3 &plusmn, 3.9%) on cell-based assays. Considering that the activity of both the UPP and ALP pathways decreases with aging, our results suggest that the hydroxybenzoic acid scaffold could be considered as a promising candidate for the development of novel modulators of the proteostasis network, and likely of anti-aging agents.

Fomitopsis officinalis is a medicinal mushroom used in traditional European eighteenth and nineteenth century folk medicine. Fruiting bodies of F. officinalis were collected from the natural environment of Świętokrzyskie Province with the consent of the General Director for Environmental Protection in Warsaw. Mycelial cultures were obtained from fragments of F. officinalis fruiting bodies. The taxonomic position of the mushroom mycelium was confirmed using the PCR method. The presence of organic compounds was determined by HPLC–DAD analysis. Bioelements were determined by AF-AAS. The biochemical composition of the tested mushroom material was confirmed with the FTIR method. Antioxidant properties were determined using the DPPH method, and the antiproliferative activity was assessed with the use of the MTT test. The presence of indole compounds (l-tryptophan, 6-methyl-d,l-tryptophan, melatonin, 5-hydroxy-l-tryptophan), phenolic compounds (p-hydroxybenzoic acid, gallic acid, catechin, phenylalanine), and sterols (ergosterol, ergosterol peroxide) as well as trace elements was confirmed in the mycelium and fruiting bodies of F. officinalis. Importantly, a high level of 5-hydroxy-l-tryptophan in in vitro mycelium cultures (517.99 mg/100 g d.w) was recorded for the first time. The tested mushroom extracts also showed antioxidant and antiproliferative effects on the A549 lung cancer cell line, the DU145 prostate cancer cell line, and the A375 melanoma cell line.

Fungal glucuronoyl esterases (FGEs) catalyze cleavage of the ester bond connecting a lignin alcohol to the xylan-bound 4-O-methyl-D-glucuronic acid of glucuronoxylans. Thus, FGEs are capable of degrading lignin-carbohydrate complexes and have potential for biotechnological applications toward woody biomass utilization. Therefore, identification and characterization of new FGEs are of critical importance. Firstly, in this study, we built a phylogenetic tree from almost 400 putative FGEs obtained on BLAST analysis and defined six main clades. In the phylogenetic tree, all the putative FGEs of ascomycetes cluster in clades I to IV, and most of the putative FGEs of basidiomycetes (B-FGEs) cluster in clades V to VI. Interestingly, several B-FGEs were found to cluster in clade II; most FGEs of clade II were found to have higher theoretical isoelectric points than those in the other five clades. To gain an insight into the putative FGEs in the clades that have not been characterized yet, we chose the FGEs of Ceriporiopsis subvermispora (CsGE) and Pleurotus eryngii (PeGE), which belong to clades V and II, respectively. The catalytic domains of both CsGE and PeGE were successfully expressed using Pichia pastoris, and then purified. Benzyl glucuronic acid was used as a substrate to confirm the activities of the CsGE and PeGE, and the hydrolyzed product, glucuronic acid, was quantified spectrophotometrically. Both CsGE and PeGE clearly exhibited the esterase activity. Additionally, we demonstrated that PeGE exhibits high tolerance toward several denaturing agents, which may make it a potentially more applicable enzyme.

Brown-rot fungi preferentially degrade softwood and cause severe breakdown of wooden structures. At the initial stage of the brown-rot decay, penetrating hyphae of the fungi are observed in ray parenchyma. Since starch grains are known to be present in the ray parenchyma of sapwood, investigation of the functions and roles of the starch-degrading enzymes is important to understand the initial stage of brown-rot decay. We purified and characterized two starch-degrading enzymes, an α-amylase (FpAmy13A) and a glucoamylase (FpGLA15A), from the brown-rot fungus, Fomitopsis palustris, and cloned the corresponding genes. The optimal temperature for both enzymes was 60 °C. FpAmy13A showed higher activity at a broad range of pH from 2.0 to 5.0, whereas FpGLA15A was most active at pH 5.0–6.0. Notable thermal stability was found for FpGLA15A. Approximately 25% of the activity remained even after treatment at 100 °C for 30 min in sodium phosphate buffer at pH 7.0. These different characteristics imply the different roles of these enzymes in the starch degradation of wood.

Glutathione S-transferases (GSTs) of wood-degrading fungi play essential roles in cellular detoxification processes and endogenous metabolism. Fungal GSTs of GSTFuA class are suggested to be involved in lignin degradation. Ceriporiopsis subvermispora is one of the important model fungi of the selective lignin degraders, we found it interesting to study its GSTs. Here, we characterized the activities of two GSTs of the GSTFuA class of C. subvermispora (CsGST63524 and CsGST83044). A high-yield expression systems involving Escherichia coli was developed for each of these enzymes. Both enzymes were found to exhibit GSH-conjugation activity toward 1-chloro-2,4-dinitrobenzene, and GSH-peroxidase activity toward cumene hydroperoxide. Both enzymes showed high GSH-conjugation activity under basic conditions (pH8.0 to 9.0), and the optimum temperature for their activity was 40°C. In addition, three fluorescent compounds were used i.e., methylumbelliferyl acetovanillone was used to monitor etherase activity, and 5-chloromethylfluorescein diacetate and 4-methylumbelliferyl acetate to monitor esterase activity. CsGST83044 exhibited both etherase and esterase activities, while CsGST63524 displayed only esterase activity, which was much higher than that of CsGST83044. These findings imply the functional diversity of the GSTFuA class GSTs of C. subvermispora, suggesting that each protein plays distinctive roles in both the fungal detoxification system and wood compound metabolism.

A glycosidase of the basidiomycete Bjerkandera adusta (BadGluc) was found in screenings to possess a strong decolorizing ability towards malvidin-3-galactoside, an anthocyanin abundant in various berry fruits. The BadGluc was purified from the culture supernatant via FPLC, and the corresponding gene was identified which showed low similarity to other characterized glucosidases. Scanning the primary sequence with PROSITE no active site motif was detected. Eventually, a specific 18 aa consensus pattern was identified manually. The active site motif possessed an undescribed sequence which was only found in a few hypothetical proteins. The corresponding gene was cloned and expressed in Pichia pastoris GS115 yielding activities up to 100 U/L using 4-nitrophenyl-β-d-glucopyranoside (pNPG) as substrate. The enzyme possessed a good temperature (70% after 1 h at 50°C) and pH stability (70% between pH 2 and 7.5), and preferably catalysed the hydrolysis of delphinidin-3-glucoside and cyanidin-3-glucoside, regardless of the position of the terminal Hexa-His tag. This novel glucosidase worked in aqueous solution as well as on pre-stained fabrics making it the first known candidate anthocyanase for applications in the detergent and food industries.

A gene encoding an endoglucanase belonging to subfamily C of glycoside hydrolase family 45 (GH45) was identified in the brown rot fungus Fomitopsis palustris and functionally expressed in Pichia pastoris. The recombinant protein displayed hydrolytic activities toward various substrates such as carboxymethyl cellulose, phosphoric acid swollen cellulose, glucomannan, lichenan, and β-glucan. In particular, the enzyme had a unique catalytic efficiency on β-1,4-glucans rather than mixed β-1,3/1,4-glucans as compared to other GH45 endoglucanases. The fungal enzyme was relatively thermostable, retaining more than 91.4% activity at 80 °C for 1 h. Site-directed mutagenesis studies revealed that the mutants N95D and D117N had significantly reduced enzymatic activities, indicating that both residues are essential for the catalytic reaction. Our study expands knowledge and understanding of the catalytic mechanism of GH45 subfamily C enzymes and also suggests that this thermostable endoglucanase from F. palustris has great potential in industrial applications.

Short-chain fatty acids (SCFAs), which are the end products of carbohydrate fermentation in the gut, mainly contribute to energy metabolism in mammals. The amount of SCFAs produced during fermentation is an important parameter that characterizes the fermentation capacity of a system. This paper reports on the fermentation characteristics of exopolysaccharides (EPS) isolated from Fomitopsis castaneus Imaz, a wood-rot fungal species. We isolated and purified the main EPS fraction by freeze drying and DEAE-Sepharose fast flow chromatography. We then analyzed the monosaccharide composition of EPS. The isolated EPS was mainly composed of glucose, galactose, rhamnose, mannose, and arabinose. The characteristic absorption peaks of sugar esters were also detected. Fresh fecal extracts from healthy adults and children were used as fermentation substrate to simulate the human intestinal environment (anaerobic conditions at 37°C) and study the fermentation characteristics of the purified EPS. Adding the isolated EPS to the fermentation system of the simulated intestinal environment increased the SCFAs content in the fecal extract of adults and children. However, the yield of SCFAs, particularly butyric acid, in the fermentation system of fecal extract in children was higher than that in adults. Furthermore, adding exogenous lactic acid bacteria, such as Enterococcus fecalis and Enterococcus fecium, to the fermentation system effectively increased the SCFAs concentration in the model intestinal system of the children. By contrast, adding E. fecalis, Lactobacillus rhamnosus, and E. fecium increased the content of the produced SCFAs in the system of adults. Those results indicate that EPS isolated from F. castaneus Imaz was effectively fermented in the simulated intestinal environments, and the fermentation capability was enhanced by adding microbial flora.

Chemical investigation of a bioactive methanolic extract of the fruiting bodies of Piptoporus betulinus led to the isolation of five previously undescribed lanostane triterpenoids named piptolinic acids A−E, as well as five known lanostane triterpenoids. Their structures were elucidated on the basis of 1D and 2D NMR spectroscopic and HRESIMS analysis. Piptolinic acid A with an unusual moiety (3-hydroxy-4-methoxycarbonyl-3-methylbutyryloxy) at C-3 exhibited comparable cytotoxic activity against human promyelocytic leukemia cell line HL-60 (IC 50 = 1.77 μM) and human acute monocytic leukemia cell line THP-1 (IC 50 = 8.21 μM) to those of positive control, fluorouracil (IC 50 = 6.38 and 4.41 μM, respectively).

The functions and properties of fungal sheath, an extracellular polysaccharide produced by many white-rot fungi, have been studied. However, the strong adherence of the sheath to fungal hyphae had been a major impediment in preparing intracellular proteins from the fungi and analyzing their cellular responses. To overcome this issue, we developed a rapid and easy method to remove the polysaccharide sheath using a selective lignin degrader, Ceriporiopsis subvermispora, which produces large sheath amounts in the presence of a lignin-derived aromatic compound. Using this approach, we achieved thorough removal of sheath and cell disruption using beads and a solution with a high protein-solubilizing power, which enabled the efficient extraction of intracellular proteins from C. subvermispora surrounded by sheath. In addition, for proteomic analysis, we investigated whether these extracted proteins were compatible with two-dimensional electrophoresis. By efficiently concentrating on protein solubilization in the first dimension and using a stacking gel in the second dimension, we successfully obtained a high-resolution proteome map of C. subvermispora. We also used the same proteins for fluorescence two-dimensional difference gel electrophoresis to obtain the quantitative protein expression profiles. These steps demonstrated that two-dimensional electrophoresis-based proteomics can be used to clarify the composition of intracellular proteins from sheath-producing white-rot fungi.

In this work we communicate the heterologous expression of a laccase from Coriolopsis gallica in Pichia pastoris . This enzyme has been reported to efficiently degrade a variety of pollutants such as industrial dyes. The expression strategy included using a previously reported modified α-factor preproleader for enhanced secretion and p AOX1 , a methanol-responsive promoter. Methanol concentration, copper salts concentration and temperature were varied in order to enhance laccase expression in this heterologous system. A volumetric activity of 250 U/L was achieved after 12-day culture in Fernbach flasks. The protein was recovered from the supernatant and purified, obtaining a preparation with 90% electrophoretic purity. The catalytic constants of the recombinant enzyme are almost identical to the fungal enzyme, thus rendering this system a useful tool for protein engineering of laccase from C. gallica .

In this work, a novel polysaccharide (named DBFM3) was isolated from mycelia of Bjerkandera fumosa by DEAE-32 and Sepharose CL–6 B column chromatography. High-performance gel permeation chromatography showed that DBFM3 was homogeneous, with an average molecular weight of 1.8 × 105 Da. Structural characteristics of the purified fraction were investigated by high-performance liquid chromatography (HPLC), FT-IR, and NMR. HPLC analysis indicated that DBFM3 was composed of mannose, galacturonic acid, and galactose at molar ratios of 1:18.16:0.702. Spectral analysis suggested that DBFM3 had (1 → 6), (1 → 3,6), (1 → 3) linkages and pyranose conformation. Antioxidant assay in vitro showed that DBFM3 exhibited 1,1-diphenyl-2-picrylhydrazyl (DPPH) and hydroxyl radical scavenging activity, and has protective effect against DNA damage and damage to SH-SY5Y cells induced by H2O2. Immunological tests indicated that DBFM3 significantly increased lymphocyte proliferation in vitro. Furthermore, DBFM3 increased the proliferation of lymphocytes in the presence of concanavalin A or lipopolysaccharide as mitogen.

White-rot basidiomycete Coriolopsis gallica HTC is one of the main biodegraders of poplar. In our previous study, we have shown the strong capacity of C. gallica HTC to degrade lignocellulose. In this study, equal amounts of total RNA fromC. Gallica HTC cultures grown in different conditions were pooled together. Illumina paired-end RNA sequencing was performed, and 13.2 million 90-bp paired-end reads were generated. We chose the Merged Assembly of Oases data-set for the following blast searches and gene ontology analyses. The reads were assembled de novo into 28,034 transcripts (≥ 100 bp) using combined assembly strategy MAO. The transcripts were annotated using Blast2GO. In all, 18,810 transcripts (≥100 bp) achieved BLASTX hits, of which, 7048 transcripts had GO term and 2074 had ECs. The expression level of 11 lignocellulolytic enzyme genes from the assembled C. gallica HTC transcriptome were detected by real-time quantitative polymerase chain reaction. The results showed that expression levels of these genes were affected by carbon source and nitrogen source at the level of transcription. The current abundant transcriptome data allowed the identification of many new transcripts in C. gallica HTC. Data provided here represent the most comprehensive and integrated genomic resources for cloning and identifying genes of interest from C. gallica HTC. Characterization of C. gallica HTC transcriptome provides an effective tool to understand mechanisms underlying cellular and molecular functions of C. gallica HTC.

Wood rotting basidiomycetes produce extracellular mucilaginous sheaths interfacing fungal hyphae and plant biomass. While the versatility of these fungal sheaths has been addressed, sheaths generated by selective white-rot fungi remain poorly understood. To fill this gap, the sheath produced by the basidiomycete Ceriporiopsis subvermispora, which degrades lignin while inflicting limited cellulose damage, was analyzed in this study. Fluorescence and transmission electron microscopy revealed that the sheath formed three days after inoculation into a beech wood slice on an agar plate and was embedded at the interface between fungal hyphae and wood cell walls. The sheath's chemical structure was evaluated from fungus cultures in a liquid medium containing [U-13C6]-d-glucose and beech wood slices. Compositional analysis, methylation analysis, and 13C NMR demonstrated that the sheath mainly consisted of a comb-like β-1,6-glucopyranose residue-branched β-1,3-glucan, which is advantageous to retain water and extracellular secondary metabolites.

The aim of this study was to evaluate of possibility of biotransformation and toxicity effect of monoanthraquinone dyes in cultures of Bjerkandera adusta CCBAS 930. Phenolic compounds, free radicals, phytotoxicity (Lepidium sativum L.), ecotoxicity (Vibrio fischeri) and cytotoxicity effect were evaluated to determine the toxicity of anthraquinone dyes before and after the treatment with B. adusta CCBAS 930. More than 80% of ABBB and AB129 was removed by biodegradation (decolorization) and biosorption, but biodegradation using oxidoreductases was the main dye removing mechanism. Secondary products toxic to plants and bacteria were formed in B. adusta strain CCBAS 930 cultures, despite efficient decolorization. ABBB and AB129 metabolites increased reactive oxygen species (ROS) production in human fibroblasts, but did not increase LDH release, did not affect the resazurine reduction assay and did not change caspase-9 or caspase-3 activity.

Ceriporiopsis subvermispora (C. subvermispora), one of the white-rot fungi, is known as a selective lignin degrader of the woody biomass. Glutathione S-transferases (GSTs) are multifunctional enzymes that are capable of catalyzing the reactions involved in detoxification and metabolic pathways. In this study, a GST of C. subvermispora, named CsGST63524, was overexpressed in E. coli, and then purified by affinity, anion exchange, and size exclusion column chromatography. The crystal structures of the CsGST63524 in ligand-free and complex with GSH were refined at 2.45 and 2.50 A resolutions, respectively. The sulfur atom of glutathione forms a hydrogen bond with Ser21 of CsGST63524, indicating it is a serine-type GST. Mutagenesis of Ser21 unexpectedly indicated that this serine residue is not essential for the enzymatic activity of CsGST63524. Comparative sequence and structural analyses, together with functional mutagenesis, newly identified the enzymatically important non-canonical amino acid residues, Asn23 and Tyr45, other than the serine residue.

Wood-degrading brown rot fungi are essential recyclers of plant biomass in forest ecosystems. Their efficient cellulolytic systems, which have potential biotechnological applications, apparently depend on a combination of two mechanisms: lignocellulose oxidation (LOX) by reactive oxygen species (ROS) and polysaccharide hydrolysis by a limited set of glycoside hydrolases (GHs). Given that ROS are strongly oxidizing and nonselective, these two steps are likely segregated. A common hypothesis has been that brown rot fungi use a concentration gradient of chelated metal ions to confine ROS generation inside wood cell walls before enzymes can infiltrate. We examined an alternative: that LOX components involved in ROS production are differentially expressed by brown rot fungi ahead of GH components. We used spatial mapping to resolve a temporal sequence in Postia placenta, sectioning thin wood wafers colonized directionally. Among sections, we measured gene expression by whole-transcriptome shotgun sequencing (RNA-seq) and assayed relevant enzyme activities. We found a marked pattern of LOX up-regulation in a narrow (5-mm, 48-h) zone at the hyphal front, which included many genes likely involved in ROS generation. Up-regulation of GH5 endoglucanases and many other GHs clearly occurred later, behind the hyphal front, with the notable exceptions of two likely expansins and a GH28 pectinase. Our results support a staggered mechanism for brown rot that is controlled by differential expression rather than microenvironmental gradients. This mechanism likely results in an oxidative pretreatment of lignocellulose, possibly facilitated by expansin- and pectinase-assisted cell wall swelling, before cellulases and hemicellulases are deployed for polysaccharide depolymerization.

Steroid sulfatase (STS) transforms hormone precursors into active steroids. Thus, it represents a target of intense research regarding hormone-dependent cancers. In this study, three ligand-based pharmacophore models were developed to identify STS inhibitors from natural sources. In a pharmacophore-based virtual screening of a curated molecular TCM database, lanostane-type triterpenes (LTTs) were predicted as STS ligands. Three traditionally used polypores rich in LTTs, i.e., Ganoderma lucidum Karst., Gloeophyllum odoratum Imazeki, and Fomitopsis pinicola Karst., were selected as starting materials. Based on eighteen thereof isolated LTTs a structure activity relationship for this compound class was established with piptolinic acid D (1), pinicolic acid B (2), and ganoderol A (3) being the most pronounced and first natural product STS inhibitors with IC50 values between 10 and 16 µM. Molecular docking studies proposed crucial ligand target interactions and a prediction tool for these natural compounds correlating with experimental findings.

Versatile peroxidase (VP) from Bjerkandera adusta is an enzyme able to oxidize bulky and high-redox substrates trough a Long-Range Electron Transfer (LRET) pathway. In this study, the introduction of radical-forming aromatic amino acids by chemical modification of the protein surface was performed, and the catalytic implications of these additional surface active-sites on the oxidation of 2,6-dimethylphenol, Mn2+ and Remazol Brilliant Blue R (RBBR) were determined. These three different substrates are oxidized in different active-sites of enzyme molecule, of which the high redox RBBR the only one that is transformed by an external radical formed on the protein surface. Both catalytic constants kcat and KM were significantly affected by the chemical modifications. Tryptophan- and tyrosine-modified VP showed higher catalytic transformation than the unmodified enzyme for RBBR, while the Mn2+ oxidation was significantly reduced by all chemical modifications. Electron Paramagnetic Resonance studies demonstrated the formation of additional protein-based radicals after the chemical modification with radical-forming amino acids. In addition, the catalytic rate of the LRET-mediated transformation showed a good correlation with the ionization energy of the additional amino acid on the protein surface.

A new manganese peroxidase-producing white-rot basidiomycete fungus was isolated from symptomatic wood of the camphor trees Cinnamomum camphora (L.) at the Hamma Botanical Garden (Algeria) and identified as Trametes pubescens strain i8. The enzyme was purified (MnP TP55) to apparent electrophoretic homogeneity and biochemically characterized. The specific activity and Reinheitzahl value of the purified enzyme were 221 U/mg and 2.25, respectively. MALDI-TOF/MS analysis revealed that the purified enzyme was a monomer with a molecular mass of 55.2 kDa. The NH2-terminal sequence of the first 26 amino acid residues of MnP TP55 showed high similarity with those of white-rot fungal peroxidases. It revealed optimal activity at pH 5 and 40 °C. This peroxidase was completely inhibited by sodium azide and potassium cyanide, suggesting the presence of heme-components in its tertiary structure. Interestingly, MnP TP55 showed higher catalytic efficiency, organic solvent-tolerance, dye-decolorization ability, and detergent-compatibility than that of horseradish peroxidase (HRP) from roots of Armoracia rustanica, manganese peroxidase from Bjerkandera adusta strain CX-9 (MnP BA30), and manganese peroxidase from Phanerochaete chrysosporium (MnP PC). Overall, the findings provide strong support for the potential candidacy of MnP TP55 for environmental applications, mainly the development of enzyme-based technologies for lignin biodegradation, textile-dyes biodecolorization, and detergent formulations.

Oxidative enzymes of white-rot fungi play a key role in lignin biodegradation. Among those fungus, Ceriporiopsis subvermispora degrades lignin before cellulose in wood; C. subvermispora is the only fungus that secretes all known types of manganese peroxidases (CsMnPs). Utilization of lignin-degrading peroxidases has been limited so far due to the lack of efficient preparation methods and intensive characterization. In this study, we developed a highly efficient method to prepare active CsMnPs through soluble expression by E. coli, which had long been impossible. The genes of MnPs selected from each subfamily were codon-optimized and expressed under the control of a cold shock promoter. A proper level of heme incorporation was achieved by continuous addition of hemin during cultivation. As much as 3 mg of purified MnPs was obtained from 100 mL culture, which is an about 20-fold higher yield than that from inclusion bodies through refolding. Further improvement of the solubility on the expression was achieved by combinatorial coexpression of chaperones. All obtained MnPs had heme-to-protein ratios as high as those of native MnPs. They were all active below pH 5. Our method is applicable to other fungal-secreted enzymes should help the progress of their basic characterization and application for better utilization of woody biomass.

WOS: 000359166300017, PubMed: 25940531, Chitin immediately suggests the representatives of the kingdom Fungi, as well as such phyla as Annelida, Mollusca, Porifera, Cnidaria and, mostly, Arthropoda. Although Bryozoa also represents a chitin-containing phylum, no study has been developed yet on the isolation or characterization of the chitin from it. In this study, physiochemical properties of the chitin isolated from Plumatella repens belonging to the phylum Bryozoa was determined for the first time. The chitin structure was also studied comparatively by isolating chitin from an insect species (Palomena prasina) of the phylum Arthropoda, and Fames fomentarius belonging to the kingdom Fungi. It was observed that the bryozoan chitin was in the a form, as in the arthropod and fungal chitins. The chitin contents in the dry weight of the bryozoan, fungal and insect species were observed to be 13.3%, 2.4%, and 10.8%, respectively. The insect chitin exhibited the highest thermal stability followed by that of the bryozoan and then the fungal chitins. Surface morphologies reveal that the insect and bryozoan chitins were composed of nano fibre and pore structures, whereas the fungal chitin had no pores or fibres. The crystallinity of the insect chitin (CrI = 84.9%) was higher than the bryozoan (CrI = 60.1%) and fungal chitins (CrI = 58.5%). (C) 2015 Elsevier B.V. All rights reserved.

The variable C-terminal tail of manganese peroxidases, a group of enzymes involved in lignin degradation, is implicated in their catalytic and stability properties, as shown by new crystal structures, molecular-simulation and directed-mutagenesis data. Based on this structural–functional evaluation, short and long/extralong manganese peroxidase subfamilies have been accepted; the latter are characterized by exceptional stability, while it is shown for the first time that the former are able to oxidize other substrates at the same site where manganese(II) is oxidized., The genome of Ceriporiopsis subvermispora includes 13 manganese peroxidase (MnP) genes representative of the three subfamilies described in ligninolytic fungi, which share an Mn2+-oxidation site and have varying lengths of the C-terminal tail. Short, long and extralong MnPs were heterologously expressed and biochemically characterized, and the first structure of an extralong MnP was solved. Its C-terminal tail surrounds the haem-propionate access channel, contributing to Mn2+ oxidation by the internal propionate, but prevents the oxidation of 2,2′-azino-bis(3-ethylbenzothiazoline-6-sulfonate) (ABTS), which is only oxidized by short MnPs and by shortened-tail variants from site-directed mutagenesis. The tail, which is anchored by numerous contacts, not only affects the catalytic properties of long/extralong MnPs but is also associated with their high acidic stability. Cd2+ binds at the Mn2+-oxidation site and competitively inhibits oxidation of both Mn2+ and ABTS. Moreover, mutations blocking the haem-propionate channel prevent substrate oxidation. This agrees with molecular simulations that position ABTS at an electron-transfer distance from the haem propionates of an in silico shortened-tail form, while it cannot reach this position in the extralong MnP crystal structure. Only small differences exist between the long and the extralong MnPs, which do not justify their classification as two different subfamilies, but they significantly differ from the short MnPs, with the presence/absence of the C-terminal tail extension being implicated in these differences.

Brown rot fungi initiate wood decay using oxidative pretreatments to improve access for cellulolytic enzymes. These pretreatments are incompatible with enzymes, and we recently showed that Postia placenta overcomes this issue by delaying glycoside hydrolase (GH) gene upregulation briefly (

Two extracellular peroxidases from Bjerkandera adusta strain CX-9, namely a lignin peroxidase (called LiP BA45) and manganese peroxidase (called MnP BA30), were purified simultaneously by applying successively, ammonium sulfate precipitation-dialysis, Mono-S Sepharose anion-exchange and Sephacryl S-200 gel filtration and biochemically characterized. The sequence of their NH2-terminal amino acid residues showed high homology with those of fungi peroxidases. Matrix assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF/MS) analysis revealed that the purified enzymes MnP BA30 and LiP BA45 were a monomers with a molecular masses 30125.16 and 45221.10Da, respectively. While MnP BA30 was optimally active at pH 3 and 70°C, LiP BA45 showed optimum activity at pH 4 and 50°C. The two enzymes were inhibited by sodium azide and potassium cyanide, suggesting the presence of heme-components in their tertiary structures. The Km and Vmax for LiP BA45 toward 2,4-Dichlorolphenol (2,4-DCP) were 0.099mM and 9.12U/mg, respectively and for MnP BA30 toward 2,6-Dimethylphenol (2,6-DMP), they were 0.151mM and 18.60U/mg, respectively. Interestingly, MnP BA30 and LiP BA45 demonstrated higher catalytic efficiency than that of other tested peroxidases (MnP, LiP, HaP4, and LiP-SN) and marked organic solvent-stability and dye-decolorization efficiency. Data suggest that these peroxidases may be considered as potential candidates for future applications in distaining synthetic-dyes.

Two new polyols, (2E,4E)-octa-2,4-diene-1,6,7-triol (1) and (E)-7,8-dihydroxyoct-5-en-2-one (2) were isolated from cultures of the basidiomycete Daedaleopsis tricolor. Their structures were elucidated by spectroscopic methods including extensive 2D NMR techniques. Two compounds showed no significant activity against five human cancer cell lines.

The secretion of exopolysaccharides and oxalic acid in cultures of a white rot Ganoderma applanatum strain and a brown rot Tyromyces palustris strain were tested in terms of culture time, pH range, and temperature. The high yield of exopolysaccharides (EPS) required a moderate temperature of 28 °C for G. applanatum and 20 °C for T. palustris. G. applanatum and T. palustris accumulated more EPS when the concentration of the carbon source (maltose for G. applanatum and fructose for T. palustris) was 30 g/L. The results indicate that the production of oxalic acid by G. applanatum is correlated with the initial pH value of the culture medium and the concentration of oxalic acid increased to 1.66 ± 0.2 mM at the initial pH of 6.5 during the fungal growth. During the growth of T. palustris, the reduction of the initial pH value of the growing medium lowered the oxalic acid concentration from 7.7 ± 0.6 mM at pH 6.0 to 1.99 ± 0.2 mM at pH 3.5. T. palustris accumulated considerably more oxalic acid than G. applanatum and its presence did not affect significantly the production of exopolysaccharides. We also observed that the maximum amounts of exopolysaccharides secreted during cultivation of G. applanatum and T. palustris were 45.8 ± 1.2 and 19.1 ± 1.2 g/L, respectively.

A novel laccase was isolated from the culture filtrate of the brown-rot fungus, Fomitopsis pinicola. Enzyme production reached its highest level after cultivation for 8 days at 25°C. The enzyme was purified by ultrafiltration, ion exchange chromatography, gelfiltration chromatography, and hydrophobic interaction chromatography. Zymography analysis of the purified enzyme showed a laccase band with a molecular mass of 92 kDa. The molecular weight of the enzyme was 92 kDa as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and gel filtration chromatography. The enzyme also had an isoelectric point of 3.8. The optimum temperature and pH for enzyme activity were 80°C and 3.0, respectively. Enzyme activity was relatively stable in the pH range from 1.5 to 11.0 and at temperatures below 40°C. The N-terminal amino acid sequence of the enzyme was DTHKAEIACRFKDLG. Enzyme activity was potently inhibited by NaN3 and SDS. The enzyme showed the highest specific activity for 2,2-azino-bis(3-ethylthiazoline-6-sulfonate) (ABTS) as a substrate. The Km value of the enzyme for ABTS substrate was 0.28 mM with a Vmax value of 4.5 U/min. The enzyme degraded several recalcitrant dyes at different time intervals during decolorization. Therefore, the novel laccase from F. pinicola may be potentially useful in industry.

Membrane inlet mass spectrometry (MIMS) uses a semipermeable membrane as an inlet to a mass spectrometer for the measurement of the concentration of small uncharged molecules in solution. We report the use of MIMS to characterize the catalytic properties of oxalate oxidase (E.C. 1.2.3.4) from Ceriporiopsis subvermispora (CsOxOx). Oxalate oxidase is a manganese dependent enzyme that catalyzes the oxygen-dependent oxidation of oxalate to carbon dioxide in a reaction that is coupled with the formation of hydrogen peroxide. CsOxOx is the first bicupin enzyme identified that catalyzes this reaction. The MIMS method of measuring OxOx activity involves continuous, real-time direct detection of oxygen consumption and carbon dioxide production from the ion currents of their respective mass peaks. 13 C 2 -oxalate was used to allow for accurate detection of 13 CO 2 ( m / z 45) despite the presence of adventitious 12 CO 2 . Steady-state kinetic constants determined by MIMS are comparable to those obtained by a continuous spectrophotometric assay in which H 2 O 2 production is coupled to the horseradish peroxidase catalyzed oxidation of 2,2′-azinobis-(3-ethylbenzthiazoline-6-sulphonic acid). Furthermore, we used MIMS to determine that NO inhibits the activity of the CsOxOx with a K I of 0.58 ± 0.06 μM.

Now a day, laccases are the most promising enzymes in the area of biotechnology and synthesis. One of the best applications of laccases is the selective oxidation of aromatic methyl group to aldehyde group. Such transformations are valuable because it is difficult to stop the reaction at aldehyde stage. Chemical methods used for such biotransformations areexpensive and give poor yields. But, the laccase-catalyzed biotransformations of such type are non-expensive and yield is excellent. Authors have used crude laccase obtained from the liquid culture growth medium of fungal strain Coriolus versicolor MTCC-138 for the biotransformations of toluene, 3-nitrotoluene, and 4-chlorotoluene to benzaldehyde, 3-nitrobenzaldehyde, and 4-chlorobenzaldehyde, respectively, instead of purified laccase because purification process requires much time and cost. This communication reports that crude laccase can also be used in the place of purified laccase as effective biocatalyst.

The white-rot basidiomycetes efficiently degrade all wood cell wall polymers. Generally, these fungi simultaneously degrade cellulose and lignin, but certain organisms, such as Ceriporiopsis subvermispora , selectively remove lignin in advance of cellulose degradation. However, relatively little is known about the mechanism of selective ligninolysis. To address this issue, C. subvermispora was grown in liquid medium containing ball-milled aspen, and nano-liquid chromatography-tandem mass spectrometry was used to identify and estimate extracellular protein abundance over time. Several manganese peroxidases and an aryl alcohol oxidase, both associated with lignin degradation, were identified after 3 days of incubation. A glycoside hydrolase (GH) family 51 arabinofuranosidase was also identified after 3 days but then successively decreased in later samples. Several enzymes related to cellulose and xylan degradation, such as GH10 endoxylanase, GH5_5 endoglucanase, and GH7 cellobiohydrolase, were detected after 5 days. Peptides corresponding to potential cellulose-degrading enzymes GH12, GH45, lytic polysaccharide monooxygenase, and cellobiose dehydrogenase were most abundant after 7 days. This sequential production of enzymes provides a mechanism consistent with selective ligninolysis by C. subvermispora .

A novel strategy of enhancing laccase production by absorbent fermentation was investigated. Peanut shell was used as nutrient carrier for laccase production by Funalia trogii IFP0027 in the absorbent fermentation. The maximum laccase production was reached to 11,900 U/l, which was 4.97 times higher than that of the control group. The results indicated that carbohydrates and phenolic substances especially flavonoids contained in peanut shell stimulated laccase production by F. trogii. Meanwhile, the peanut shell nutrient carrier could not only alleviate the oxidative damage, owing to strong scavenging activity on hydroxyl, but also relieve the mechanical stresses to form small and regular microbial pellets. Therefore, the absorbent fermentation using peanut shell as nutrient carrier shows enormous potential in enhancing laccase production.

Two new ylangene-type sesquiterpenoids, postinins A (1) and B (2), were isolated from cultures of the fungus Postia sp. Structures 1 and 2 were elucidated on the basis of extensive spectroscopic analysis. The bioactivity evaluation showed that both compounds had significant inhibitory activities against protein tyrosine phosphatase 1B, and SH2-containing cytoplasmic tyrosine phosphatase-1 and -2 with IC₅₀ values of 1.6-6.2 μg/ml.

Enzymatic lignin activation may be an environmentally friendly alternative to the use of chemicals in the production of wood fibers composites. Most studies on enzymatic activation of lignin for improving the adhesion of lignocellulosic products have been carried out using laccases. In this work, the use of a versatile peroxidase (VP) from the white-rot fungus Bjerkandera sp. (anamorph R1) for activating Kraft lignin was studied. The effect of enzyme dosage, incubation time, and H(2)O(2) addition profile on lignin activation was evaluated by quantifying the phenoxy radicals formed using electron paramagnetic resonance (EPR) spectroscopy. Two alternative enzymatic systems based on the use of VP (a two-stage and an enzymatic cascade system) were also assayed. At optimal conditions (dose of 15 U g(-1) and continuous addition of H(2)O(2) (5.24 μmol h(-1)) during 1 h) the content of phenoxy radicals was doubled as compared with an untreated control. Moreover, using the two-stage VP system, a lignin activation similar to that found at optimal conditions could be reached in a shorter time.

The inhibitor and substrate specificities of versatile peroxidase from Bjerkandera fumosa (VPBF) were studied. Two different effects were found: NaN(3), Tween-80, anthracene, and fluorene decreased the activity of VPBF, but p-aminobenzoic acid increased it. A mixed mechanism of effector influence on the activity of this enzyme was shown. The catalytic properties of VPBF in the oxidation of mono- and polycyclic aromatic compounds were studied also. 2,7-Diaminofluorene, ABTS, veratryl alcohol, and syringaldazine can be oxidized by VPBF in two ways: either directly by the enzyme or by diffusible chelated Mn(3+) as an oxidizing agent. During VPBF oxidation of 2,7-diaminofluorene, both with and without Mn(2+), biphasic kinetics with apparent saturation in both micromolar and millimolar ranges were obtained. In the case of ABTS, inhibition of VPBF activity by an excess of substrate was observed. Direct oxidation of p-aminobenzoic acid by versatile peroxidase was found for the first time. The oxidation of three- and four-ring PAHs by VPBF was investigated, and the oxidation of anthracene, phenanthrene, fluorene, pyrene, chrysene, and fluoranthene was shown. The products of PAH oxidation (9,10-anthraquinone, 9,10-phenanthrenequinone, and 9-fluorenone) catalyzed by VPBF were identified.

Commercial application of laccase is often hampered by insufficient enzyme stocks, with very low yields obtained from natural sources. This study aimed to improve laccase production by mutation of a Coriolopsis gallica strain and to determine the biological properties of the mutant. The high-yield laccase strain C. gallica TCK was treated with N-methyl-N-nitro-N-nitrosoguanidine and ultraviolet light. Among the mutants isolated, T906 was found to be a high-production strain of laccases. The mutant strain T906 was stabilized via dozens of passages, and the selected ones were further processed for optimization of metallic ion, inducers, and nutritional requirements, which resulted in the optimized liquid fermentation medium MF9. The incubation temperature and pH were optimized to be 30°C and 4.5, respectively. The mutant strain T906 showed 3-times higher laccase activity than the original strain TCK under optimized conditions, and the maximum laccase production (303 U/ml) was accomplished after 13 days. The extracellular laccase isoenzyme 1 was purified and characterized from the two strains, respectively, and their cDNA sequence was determined. Of note, the laccase isoenzyme 1 transcription levels were overtly increased in T906 mycelia compared with values obtained for strain TCK. These findings provide a basis for C. gallica modification for the production of high laccase amounts.

New drimane-type sesquiterpene cryptoporol A (1), cryptoporic acid derivative 6′-cryptoporic acid E methyl ester (2), and pseudouridine derivative cryptoporine A (3), as well as a known ergosterol 5α,8α-epidioxy-22E-ergosta-6,22-dien-3β-ol (4), were isolated from a 90 % alcohol extract of the fruiting bodies of Cryptoporus volvatus. The structures of these compounds were established by spectroscopic analysis and circular dichroism. 5α,8α-epidioxy-22E-ergosta-6,22-dien-3β-ol (4) exhibited antiviral activity against porcine reproductive and respiratory syndrome virus, and all compounds showed weak antioxidant activities.

Phytochemical investigation of the methanolic extract of the fruiting bodies of Fomes officinalis led to the isolation of eight 24-methyl-lanostane triterpenes named officimalonic acids A−H, along with one known lanostane triterpene. Their structures were elucidated based on the analysis of spectroscopic data, single-crystal X-ray diffraction, and electronic circular dichroism. Officimalonic acid A represents a previously unknown triterpene type with a 24-methyl-7(8 → 9)abeo-lanostane skeleton, and all of the compounds possessed a malonate half-ester moiety at C-3. Anti-inflammatory assay revealed that officimalonic acids D, E, G, H, and fomitopsin A showed potent inhibitory effects (IC50 = 5.1–8.9 μM) on nitric oxide production in lipopolysaccharide-induced RAW264.7 cells. Officimalonic acids E, G, H showed moderate cytotoxicity against H460, HepG2 and BGC-823 human cell lines.

In vitro evaluation of total phenolic contents and antiradical activities of the lignicolous fungi Fomes fomentarius and Schizophyllum commune hydrodistilled extracts was the subject of this study. This preliminary screening included four free radical species evaluated by UV–vis (DPPH•, ABTS• and •NO) and EPR (Asc•), respectively. According to the experimental data obtained, both F. fomentarius and S. commune hydrodistilled extracts may be considered as promising sources of phenolic natural products (157 and 138 mg GAE/g d.e., respectively) and other bioactives showing good anti-DPPH (1.31 μg/mL) and anti-Asc (70.40%) radical activities, respectively, at in vitro conditions.

The separation and purification of extracellular polysaccharides from Coriolus versicolor LH1 were investigated along with their α-glucosidase inhibition properties. Three polysaccharide fractions (ePS-F2-1, ePS-F3-1, and ePS-F4-1) were separated from the culture medium of LH1 using a DEAE anion-exchange column and a Sephadex™ G-50 gel filtration column. Their chemical compositions was determined. On the basis of an α-glucosidase inhibition assay, the enzyme inhibition activities of ePS-F2-1, ePS-F3-1, and ePS-F4-1 were investigated. Among these, ePS-F4-1 had the highest enzyme inhibition effects on α-glucosidase. According to the results of the chemical component analysis, ePS-F3-1 and ePS-F4-1 are the polysaccharides which are combined with triterpenoides, and ePS-F2-1 is complexed with proteins and triterpenoides.