Rapid screening and isolation of 5-lipoxygenase inhibitors in Inonotus obliquus and mechanism of action in the treatment of asthma.

Accurate screening and targeted preparative isolation of active substances in natural medicines have long been two technical challenges in natural medicine research. This study outlines a new approach to improve the efficiency of natural product preparation, focusing on rapidly and accurately screening potential active ingredients in Inonotus obliquus as well as efficiently preparing 5-lipoxidase (5-LOX) inhibitors, to provide new ideas for the treatment of asthma with Inonotus obliquus. First, we used ultrafiltration (UF) mass spectrometry to screen for three potential inhibitors of 5-LOX in Inonotus obliquus. Subsequently, the inhibitory effect of the active ingredients screened in the UF assay on 5-LOX was verified using the molecular docking technique, and the potential role of the active compounds in Inonotus obliquus for the treatment of asthma was analyzed by network pharmacology. Finally, based on the above activity screening guidelines, we used semi-preparative liquid chromatography and consecutive high-speed countercurrent chromatography to isolate three high-purity 5-LOX inhibitors such as betulin, lanosterol, and quercetin. Obviously, through the above approach, we have seamlessly combined rapid discovery, screening, and centralized preparation of the active ingredient with molecular-level interactions between the active ingredient and the protease.

Integrated omic profiling of the medicinal mushroom Inonotus obliquus under submerged conditions.

BACKGROUND: The Inonotus obliquus mushroom, a wondrous fungus boasting edible and medicinal qualities, has been widely used as a folk medicine and shown to have many potential pharmacological secondary metabolites. The purpose of this study was to supply a global landscape of genome-based integrated omic analysis of the fungus under lab-growth conditions. RESULTS: This study presented a genome with high accuracy and completeness using the Pacbio Sequel II third-generation sequencing method. The de novo assembled fungal genome was 36.13 Mb, and contained 8352 predicted protein-coding genes, of which 365 carbohydrate-active enzyme (CAZyme)-coding genes and 19 biosynthetic gene clusters (BCGs) for secondary metabolites were identified. Comparative transcriptomic and proteomic analysis revealed a global view of differential metabolic change between seed and fermentation culture, and demonstrated positive correlations between transcription and expression levels of 157 differentially expressed genes involved in the metabolism of amino acids, fatty acids, secondary metabolites, antioxidant and immune responses. Facilitated by the widely targeted metabolomic approach, a total of 307 secondary substances were identified and quantified, with a significant increase in the production of antioxidant polyphenols. CONCLUSION: This study provided the comprehensive analysis of the fungus Inonotus obliquus, and supplied fundamental information for further screening of promising target metabolites and exploring the link between the genome and metabolites.

A new dimeric (-)-5-methylmellein from cultures of the basidiomycete Inonotus sinensis.

(-)-5-Methylmellein (1) and its new dimer (2) were isolated from cultures of the basidiomycete Inonotus sinensis. Their structures were elucidated on the basis of extensive spectroscopic methods including UV, IR, HR-EI-MS, 1D NMR and 2D NMR. The structure of Compound 2 was determined by single-crystal X-ray crystallographic analysis. Compound 2 was tested for the cytotoxicities against five human cancer cell lines.

Genome sequencing of Inonotus obliquus reveals insights into candidate genes involved in secondary metabolite biosynthesis.

BACKGROUND: Inonotus obliquus is an important edible and medicinal mushroom that was shown to have many pharmacological activities in preclinical trials, including anti-inflammatory, antitumor, immunomodulatory, and antioxidant effects. However, the biosynthesis of these pharmacological components has rarely been reported. The lack of genomic information has hindered further molecular characterization of this mushroom. RESULTS: In this study, we report the genome of I. obliquus using a combined high-throughput Illumina NovaSeq with Oxford Nanopore PromethION sequencing platform. The de novo assembled 38.18 Mb I. obliquus genome was determined to harbor 12,525 predicted protein-coding genes, with 81.83% of them having detectable sequence similarities to others available in public databases. Phylogenetic analysis revealed the close evolutionary relationship of I. obliquus with Fomitiporia mediterranea and Sanghuangporus baumii in the Hymenochaetales clade. According to the distribution of reproduction-related genes, we predict that this mushroom possesses a tetrapolar heterothallic reproductive system. The I. obliquus genome was found to encode a repertoire of enzymes involved in carbohydrate metabolism, along with 135 cytochrome P450 proteins. The genome annotation revealed genes encoding key enzymes responsible for secondary metabolite biosynthesis, such as polysaccharides, polyketides, and terpenoids. Among them, we found four polyketide synthases and 20 sesquiterpenoid synthases belonging to four more types of cyclization mechanism, as well as 13 putative biosynthesis gene clusters involved in terpenoid synthesis in I. obliquus. CONCLUSIONS: To the best of our knowledge, this is the first reported genome of I. obliquus; we discussed its genome characteristics and functional annotations in detail and predicted secondary metabolic biosynthesis-related genes, which provides genomic information for future studies on its associated molecular mechanism.

Immunomodulatory effects of Inonotus obliquus polysaccharide on splenic lymphocytes infected with Toxoplasma gondii via NF-κB and MAPKs pathways.

CONTEXT: As a medicinal and edible fungus, Inonotus obliquus has been traditionally used to prevent and treat various ailments. Inonotus obliquus polysaccharide (IOP) isolated from I. obliquus processes many biological activities, our series of in vivo studies have shown that IOP protects against Toxoplasma gondii infection. OBJECTIVE: This study aimed to investigate the in vitro immunomodulatory effects and its mechanisms of IOP on mouse splenic lymphocytes infected with T. gondii. MATERIALS AND METHODS: Mouse splenic lymphocytes were infected with T. gondii tachyzoites, and treated with different concentrations of IOP. The levels of cytokines and chemokines were measured by enzyme-linked immunosorbent assay (ELISA) and reverse transcription-polymerase chain reaction (RT-PCR). The expression of toll-like receptor 2 (TLR2) and TLR4, and the modulation of nuclear factor-kappa B (NF-κB) and mitogen-activated protein kinases (MAPKs) signaling pathways were determined by Western blot. RESULTS: IOP significantly decreased the over-release of cytokine interleukin-1 beta (IL-1ß), IL-4, IL-6, interferon-gamma (IFN-γ), and tumor necrosis factor-alpha (TNF-α) in supernatant from T. gondii-infected splenic lymphocytes. IOP also effectively inhibited the overexpression of cytokines and chemokine macrophage inflammatory protein-1 (MIP-1) and monocyte chemoattractant protein-1 (MCP-1) mRNA. Furthermore, IOP down-regulated TLR2 and TLR4 expressions and inhibited the over-phosphorylation of NF-κB p65 and inhibitor κBα (IκBα) in NF-κB signaling pathway and p38, c-Jun N-terminal kinase (JNK) in MAPKs signaling pathway. By observing the effect of IOP on TNF-α secretion after pretreatment with specific inhibitors, it was further confirmed that IOP was involved in the regulation of NF-κB, p38, and JNK signaling pathways. CONCLUSIONS: These data indicate that IOP can inhibit the excessive inflammatory response caused by T. gondii infection through modulating NF-κB, p38, and JNK signaling pathways, and thus plays the in vitro anti-T. gondii role.

Biological Synthesis of Bioactive Gold Nanoparticles from Inonotus obliquus for Dual Chemo-Photothermal Effects against Human Brain Cancer Cells.

The possibility for an ecologically friendly and simple production of gold nanoparticles (AuNPs) with Chaga mushroom (Inonotus obliquus) (Ch-AuNPs) is presented in this study. Chaga extract's reducing potential was evaluated at varied concentrations and temperatures. The nanoparticles synthesized were all under 20 nm in size, as measured by TEM, which is a commendable result for a spontaneous synthesis method utilizing a biological source. The Ch-AuNPs showed anti-cancer chemotherapeutic effects on human brain cancer cells which is attributed to the biofunctionalization of the AuNPs with Chaga bioactive components during the synthesis process. Further, the photothermal ablation capability of the as-prepared gold nanoparticles on human brain cancer cells was investigated. It was found that the NIR-laser induced thermal ablation of cancer cells was effective in eliminating over 80% of the cells. This research projects the Ch-AuNPs as promising, dual modal (chemo-photothermal) therapeutic candidates for anti-cancer applications.

Supercritical CO2 Extraction of Triterpenoids from Chaga Sterile Conk of Inonotus obliquus.

Triterpenoids are among the bioactive components of Chaga, the sterile conk of the medicinal fungus Inonotus obliquus. Supercritical fluid extraction of Chaga triterpenoids was carried out with supercritical CO2, while a modified Folch method was used as a comparison. Three temperature-pressure combinations were tested varying between 314-324 K (40-50 °C) and 281-350 bars, using time- and volume-limited extractions. Six triterpenoids were identified with GC-MS and quantified with GC-FID: ergosterol, lanosterol, ß-sitosterol, stigmastanol, betulin, and inotodiol. The Folch extraction resulted in recovery of trametenolic acid, which was not extracted by supercritical CO2. Inotodiol was the major triterpenoid of all the extracts, with a yield of 87-101 mg/100 g and 139 mg/100 g, for SFEs and the Folch method, respectively. The contents of other major triterpenoids, lanosterol and ergosterol, varied in the ranges 59-63 mg/100 g and 17-18 mg/100 g by SFE, respectively. With the Folch method, the yields were 81 mg/100 g and 40 mg/100 g, respectively. The highest recovery of triterpenoids with SFE in relation to Folch was 56% and it was obtained at 324 K (50 °C) and 350 bar, regardless of extraction time or volume of CO2. The recoveries of lanosterol and stigmastanol were unaffected by SFE conditions. Despite the lower yield, SFE showed several advantages including shorter extraction time and less impact on the environment. This work could be a starting point for further studies on green extraction methods of bioactive triterpenoids from Chaga.

Evaluation of Toxicity and Efficacy of Inotodiol as an Anti-Inflammatory Agent Using Animal Model.

Chaga mushroom (Inonotus obliquus) comprises polyphenolic compounds, triterpenoids, polysaccharides, and sterols. Among the triterpenoid components, inotodiol has been broadly examined because of its various biological activities. The purpose of this study is to examine inotodiol from a safety point of view and to present the potential possibilities of inotodiol for medical usage. From chaga mushroom extract, crude inotodiol (INO20) and pure inotodiol (INO95) were produced. Mice were treated with either INO20 or INO95 once daily using oral administration for repeated dose toxicity evaluation. Serum biochemistry parameters were analyzed, and the level of pro-inflammatory cytokines in the serum was quantified. In parallel, the effect of inotodiol on food allergic symptoms was investigated. Repeated administration of inotodiol did not show any mortality or abnormalities in organs. In food allergy studies, the symptoms of diarrhea were ameliorated by administration with INO95 and INO20. Furthermore, the level of MCPT-1 decreased by treatment with inotodiol. In this study, we demonstrated for the first time that inotodiol does not cause any detrimental effect by showing anti-allergic activities in vivo by inhibiting mast cell function. Our data highlight the potential to use inotodiol as an immune modulator for diseases related to inflammation.

Effects of Compound Elicitors on the Biosynthesis of Triterpenoids and Activity of Defense Enzymes from Inonotus hispidus (Basidiomycetes).

Inonotus hispidus has various health-promoting activities, such as anticancer effects and immune-stimulating activity. The commercialization of valuable plant triterpenoids faces major challenges, including low abundance in natural hosts and costly downstream purification procedures. In this work, orthogonal design was used to compound methyl jasmonate (MeJA), salicylic acid (SA), oleic acid, and Cu2+, and the effects of combinations on the total triterpenes biosynthesized were studied. The optimal combination was screened out and its effect on the activity of PAL, CAT, and SOD was studied. The optimal concentration of oleic acid was 2% when MeJA was 100 mol/L, and the total triterpenoid content and mycelia production were 3.918 g and 85.17 mg/g, respectively. MeJA treatment induced oxidative stress, and at the same time increased the activity of related defense enzymes. Oleic acid is thought to regulate cell permeability by recombining cell membranes. It promotes the material exchange process between cells and the environment without affecting cell growth. When oleic acid was used in combination with MeJA, a synergistic effect on triterpene production was observed. In conclusion, our findings provide a strategy for triterpenoid enrichment of I. hispidus.

Influence of selenium on the mycelia of the shaggy bracket fungus, Inonotus hispidus.

BACKGROUND: Selenium (Se) is a needed trace element for animals and humans. Many fungi have effective mechanisms to acquire, transform and accumulate Se in organic form. In this study, the effects of inorganic Se (sodium selenite) on the medicinal fungus Inonotus hispidus was investigated. RESULTS: Inonotus hispidus was capable of tolerating up to 3.85 mmol L-1 selenite, at which ~85% growth inhibition was seen, with 50% growth inhibition occurring at ~1 mmol L-1 selenite. Growth in 0.29 mmol L-1 Se resulted in I. hispidus mycelium with 115 times higher Se levels compared to growth in standard media, and an organic Se content of 86% to total Se content. The influence of Se accumulation on morphological features of I. hispidus were examined by microscopic and scanning electron microscopic observation. These data revealed significant shrinkage and deformations of I. hispidus hyphae with decreased branching and collapse of clamp connections under higher Se stress. However, conidial production in I. hispidus increased dramatically. The influence of Se on mycelial growth could be recovered by reinoculation in standard media. Se accumulation had only minimal impacts on the yield of the potential selenocompounds such as amino acids, proteins and polysaccharides. By contrast, Se-enriched I. hispidus mycelium was of higher quality due to reduction in crude fat and total ash contents. CONCLUSIONS: These data provide basic and applied information on the feasibility of producing selenized I. hispidus as an enriched and better quality product. © 2021 Society of Chemical Industry.

Improving crude protein and methionine production, selective lignin degradation and digestibility of wheat straw by Inonotus obliquus using response surface methodology.

BACKGROUND: To date, fungus-assisted pretreatment of agricultural residue has not become the preferred method to produce protein-enriched and ruminally digestible animal feed because of low time efficiency of fungal delignification and protein production, i.e. the long solid-state fermentation period, and because of laccase as a potential inhibitor of cellulose activity. In this study, response surface methodology was employed to optimize the parameters in the process of producing nutritious animal feed from wheat straw with Inonotus obliquus pretreatment. RESULTS: The mineral salt solution containing (w/v) (NH4 )2 SO4 1%, MgSO4 ·7H2 O 0.03%, KH2 PO4 0.011%, Tween-80 0.4%, and corn starch 10% with pH of 7.4 was optimized. Inonotus obliquus rapidly and completely colonized on wheat straw with an ergosterol content of 280 µg g-1 dry matter, consuming 45% of lignin after 15 days of fermentation, producing maximums of lignin peroxidase (1729 IU g-1 ), manganese peroxidase (610 IU g-1 ) and laccase (98 IU g-1 ) on days 5, 15, and 25, respectively. The crude protein (102.4 g kg-1 ) of 15-day fermented wheat straw increased by ~132%. After hydrolysis, the essential protein-bound amino acids (15.3 g kg-1 ) increased by ~47%, within which Met and Lys measured ~1070% and ~60% higher. The treatment with I. obliquus also improved the in vitro gas production after 72 h (IVGP72 ) of wheat straw to 178.8 mL g-1 organic matter (~43% increase). CONCLUSION: For the first time, we found that I. obliquus is an effective white rot fungus turning wheat straw into ruminally digestible animal feed without laccase inhibitor.

Effects of supplementation of sea buckthorn press cake on mycelium growth and polysaccharides of Inonotus obliquus in submerged cultivation.

AIMS: Investigation of the influence of cultivation time and sea buckthorn press cake (Hippophaë rhamnoides) dosage on mycelium yield of Inonotus obliquus in submerged cultivation and on the yield, monomer composition, and macromolecular properties of the exopolysaccharides (EPS) from culture media and intracellular polysaccharides (IPS) extracted from mycelia. METHODS AND RESULTS: Supplementation at 5 g l-1 combined with cultivation time of 250 h granted highest yield increase in mycelia (by 122%). The supplementation reduced extraction yield and decreased the molecular weight of the main IPS population. The supplementation increased production and molecular weight of EPS. The relative content of arabinose and rhamnose in EPS positively correlated with dosage of the press cake. The press cake supplementation increased the content of galacturonic acid in IPS, but not in EPS. CONCLUSION: Sea buckthorn press cake is a food industry fibrous side stream with high oil content. It increases the cultivation yield of Inonotus obliquus mycelium and influences the produced polysaccharides. SIGNIFICANCE AND IMPACT OF THE STUDY: Mycelium is a resource of bioactive polysaccharides, attracting the interest of nutraceutical companies. Sea buckthorn press cake is a promising supplement for increasing mycelium production. The utilization of this agricultural side stream would therefore favour circular economy.

Development of ultrasound-assisted centrifugal extraction combined with two countercurrent chromatography systems for the simultaneous extraction and isolation of phytochemicals.

We proposed a method for the extraction of medicinal herbs, called ultrasound-assisted centrifugal extraction, and an online solvent concentration method. These techniques were coupled with two countercurrent chromatography systems and applied to the continuous extraction and online isolation of chemical constituents from Inonotus obliquus. Raw plants were extracted using a two-phase petroleum-ethanol-water (2.0:1.0:2.0, v/v/v) process, and then the aqueous and organic phases were concentrated using the proposed online solvent concentrator. The countercurrent chromatography preparation prior to separation includes pumping of the two-phase solution, rotating column, and equilibrium column. Following online concentration, the extracted solution was pumped into a second countercurrent chromatography process for separation. During separation, the extraction solution and concentrated extract were prepared automatically. Upon completion of the first cycle of ultrasound-assisted centrifugal extraction/two countercurrent chromatography, the second cycle experiment starts. This process can be indefinitely repeated. In this study, six target compounds with purities above 97.71% were successfully extracted and isolated online using a two-phase solvent system consisting of n-hexane-ethyl acetate-acetonitrile (4.5:1.5:5.5, v/v/v) and n-hexane-ethyl acetate-methanol-water (0.4:3.0:1.5:2.5, v/v/v/v). Compared to conventional extraction methods, the instrumental setup of the proposed method provides enhanced automation, efficiency, purity, and systematic extraction and isolation of natural products.

Antimicrobial, Antioxidant and Antiproliferative Secondary Metabolites from Inonotus nidus-pici.

Inonotus nidus-pici is a sterile conk which produces macrofungus, a neglected Central-Eastern European relative of the prized Inonotus obliquus, also known as chaga. Investigation of the methanol extract of the poroid fungus I. nidus-pici resulted in the isolation of citropremide (1), 3,4-dihydroxybenzalacetone (2) , lanosterol (3), ergost-6,8,22-trien-3ß-ol (4), and ergosterol peroxide (5). The structures of fungal compounds were determined on the basis of one- and two-dimensional NMR and MS spectroscopic analysis. Compounds 1-2 and 4-5 were evaluated for their antioxidant and antimicrobial properties against several bacterial and fungal strains. 3,4-dihydroxybenzalacetone (2) and ergost-6,8,22-trien-3ß-ol (4) demonstrated moderate antimicrobial activity, while the former possessed notable antioxidant activity in DPPH assay. The antiproliferative examinations performed on three human cancer (MES-SA, MES-SA/Dx5, A431) cell lines demonstrated that compounds 4 and 5 have notable cytotoxic activity with IC values in micromolar range. The current study represents the first report on the chemical profile of I. nidus-pici, providing a comprehensive study on the isolation and structure determination of bioactive secondary metabolites of this macrofungus.

Total Triterpenoid Extraction from Inonotus Obliquus Using Ionic Liquids and Separation of Potential Lactate Dehydrogenase Inhibitors via Ultrafiltration High-Speed Countercurrent Chromatography.

Extracts of the fungus Inonotus obliquus exhibit cytotoxic properties against different cancers; hence, this fungal species has been extensively studied. This study aimed to extract total triterpenoids from Inonotus obliquus using ionic liquids (ILs) and separate potential lactate dehydrogenase (LDH) inhibitors via ultrafiltration (UF)-high-speed countercurrent chromatography (HSCCC). Total triterpenoids from Inonotus obliquus were extracted by performing a single-factor experiment and employing a central composite design via ultrasonic-assisted extraction (UAE) and heat-assisted extraction (HAE). The extract was composed of 1-butyl-3-methylimidazolium bromide as the IL and methanol as the dispersant. Ultrafiltration-liquid chromatography (UF-LC) was used to rapidly scan the LDH inhibitors and betulin and lanosterol were identified as potential inhibitors. To obtain these target compounds, betulin and lanosterol with the purities of 95.9% and 97.8% were isolated from HSCCC within 120 min. Their structures were identified using several techniques, among which IL-HAE was fast and effective. This study reports the extraction of triterpenoids from Inonotus obliquus by IL for the first time. Collectively, the findings demonstrate that UF-LC is an effective tool for screening potential LDH inhibitors from crude extracts of I. obliquus and may help to identify bioactive substances against myocardial infarction, whereas high-purity compounds can be separated via UF-HSCCC.

Optimization for the extraction of polyphenols from Inonotus obliquus and its antioxidation activity.

In order to study the extraction process and antioxidative activity of Inonotus obliquus polyphenols (IOP), the optimal extraction process was determined by orthogonal experiment optimization. The clearance rate of DPPH and hydroxyl radicals were used as indicators to evaluate the antioxidant activity of IOP. The results showed that the optimum extraction conditions were as follows: ethanol concentration of 50%, solid-liquid ratio of 1:20, temperature of 60 °C, and 90 min. Under these conditions, the extraction yield of IOP was 2.84%. The antioxidant capacity of extracts appeared to be IOP dose-dependent, while it also presented stronger ferric reducing antioxidant power (FRAP). High Performance Liquid Chromatography (HPLC-MS) analysis indicated that the major identified polyphenol compounds extracted at the optimal conditions were ten compounds (procyanidin, caffeic acid, p-coumaric acid, isorhamnetin-3-O-glucoside, astilbin, tangeretin, gallic acid, kaempferol, quercetin, and catechin 7-xyloside). These findings indicate that I. obliquus polyphenols have the potential to be developed as a natural antioxidant and have a good application prospect.

Production of phenolic compounds and antioxidant activity via bioconversion of wheat straw by Inonotus obliquus under submerged fermentation with the aid of a surfactant.

BACKGROUND: This study investigated the effect of surfactants on wheat straw biodegradation and the growth-associated generation of exo- and endo-phenolic compounds (EPC and IPC) and antioxidant activity expression by liquid-cultured Inonotus obliquus, an edible and medicinal mushroom, also known as a white rot fungus. Changes in the chemical composition and multiscale structure of wheat straw, in the production and activity of EPC and IPC and in individual flavonoids were analyzed. RESULTS: Fungal pretreatment decreased significantly the contents of all lignocellulose components, increased and enlarged substrate porosity and caused changes in the structure of wheat straw with the aid of Triton X-100. A gradual increase in EPC and IPC production was observed up to 6.4- and 1.5-fold for 9 days. The EPC obtained on day 9 showed the highest antioxidant activity (IC50 of 30.96 mg L-1 ) against 2,2-diphenyl-1-picrylhydrazyl radicals. High-performance liquid chromatographic results indicated the presence of high amounts of epicatechin-3-gallate (ECG; (374.9 mg g-1 ) and epigallocatechin-3-gallate (EGCG; 447.2 mg g-1 ) in the EPC; other polyphenols were also enhanced but to a lesser extent. Surfactant supplementation was effective in enhancing flavonoid production and in increasing antioxidant activity in EPC. CONCLUSIONS: The results indicated enhanced accumulation of phenolic compounds, particularly ECG and EGCG in Inonotus obliquus via biodegradation and bioconversion of lignocellulose residues. They also indicated enhancement in the production of several flavonoids and also an increase in antioxidant activity in the product by a surfactant-treated process, which may be a useful way of exploiting underused lignocellulosic residues to various high-added-value functional ingredients. © 2020 Society of Chemical Industry.

Protective effect of Phaeoporus obliquus polysaccharide against acute liver injury induced by carbon tetrachloride and alcohol in mice.

Studied the optimum extraction process of polysaccharide from Phaeoporus obliquus and the effect of Phaeoporus obliquus polysaccharide on carbon tetrachloride (CCl4)- or alcohol-induced acute liver injury in mice. The main factor in influencing the extraction rate of Phaeoporus obliquus polysaccharide were extraction power and time, which was a kind of pyran glucose by infrared spectroscopy. CCl4 and alcohol were employed respectively to establish CCl4 and alcohol-induced acute liver injury mouse models. Compared with model groups mice, Phaeoporus obliquus polysaccharide treatment at the doses of 100mg/kg and 200mg/kg exhibited an obvious reduction liver index, ALP, ALT, AST levels, MDA content and TNF-α level (p<0.01) and SOD activity was increased, which was in a dose-dependent manner. Compared with the model group, the necrosis degree of hepatocytes was obviously reduced and the small fat droplets were formed in some cytoplasm, especially in high dose group, which the liver cells recovered to the level of normal group. Rt-PCR results showed that the expression of CYP2E1 mRNA in liver tissues of Phaeoporus obliquus polysaccharide groups were significantly reduced, and the difference were statistically significant compared with the model group (p<0.05). These results demonstrated that Phaeoporus obliquus polysaccharide has significantly hepatoprotective effect on CCl4 and alcohol-induced acute liver injury in mice.

Highly modified steroids from Inonotus obliquus.

Six undescribed steroids were isolated from the fungus Inonotus obliquus. Notably, compounds 1 and 2 represented the first example of 8,14-seco-4-methylpregnane. By spectroscopic data analyses, quantum chemical calculations and DP4+ probability analysis, their structures were unambiguously determined. The absolute configurations of the compounds 1-6 were defined by comparison of their experimental and calculated electronic circular dichroism (ECD) spectra data. The structure of compound 1 was also confirmed by single-crystal X-ray diffraction. All isolated steroids were evaluated for their neuroprotective activities against H2O2-induced cell injury in human neuroblastoma SH-SY5Y cells, among which compound 2 showed moderate activity at 12.5 µM.

Flavan derivative enantiomers and drimane sesquiterpene lactones from the Inonotus obliquus with neuroprotective effects.

A pair of new flavan derivative enantiomers (1a and 1b), four new drimane sesquiterpene lactones (2-5) and a known compound (6) were isolated from Inonotus obliquus. Their structures were determined by comprehensive spectroscopic analyses. Compounds 1a/1b were separated by chiral chromatographic column successfully. The absolute configurations of compounds 1-5 were determined by comparison of their experimental and calculated ECD spectra. Notably, compounds 1a/1b represented the first pair of enantiomers in nature with tetrahydrofuran-flavan skeleton. A plausible biosynthetic pathway for (±)-1 was also proposed. All isolates were evaluated for their neuroprotective activities against H2O2-induced cell injury in SH-SY5Y cells. The results showed that compound 2 exhibited moderate neuroprotective activity at the concentration of 25.0 µM.