Antifungal Activity of Cedrol from Cunninghamia lanceolate var. konishii against Phellinus noxius and Its Mechanism.

Phellinus noxius is a highly destructive fungus that causes brown root disease in trees, leading to decay and death. In Taiwan, five prized woods-Taiwania cryptomerioides, Calocedrus macrolepis var. formosana, Cunninghamia lanceolata var. konishii, Chamaecyparis formosensis, and Chamaecyparis obtusa var. formosana-are known for their fragrance and durability. This study aims to explore the anti-brown-root-rot-fungus activity of Cunninghamia lanceolata var. konishii (CL) essential oil (CLOL) and its primary components, while also delving into their mechanisms of action and inhibition pathways. The essential oil (CLOL) from CL wood demonstrated significant efficacy against P. noxius, with an inhibitory concentration (IC50) of 37.5 µg/mL. Cedrol, the major component (78.48%) in CLOL, emerged as a potent antifungal agent, surpassing the reference drug triflumizole. Further assays with cedrol revealed a stronger anti-brown-root-disease activity (IC50 = 15.7 µg/mL) than triflumizole (IC50 = 32.1 µg/mL). Scanning electron microscopy showed deformation and rupture of fungal hyphae treated with CLOL and cedrol, indicating damage to the fungal cell membrane. Cedrol-induced oxidative stress in P. noxius was evidenced by increased reactive oxygen species (ROS) levels, leading to DNA fragmentation, mitochondrial membrane potential reduction, and fungal apoptosis through the mitochondrial pathway. Gel electrophoresis confirmed cedrol-induced DNA fragmentation, whereas TUNEL staining demonstrated increased apoptosis with rising cedrol concentrations. Moreover, protein expression analysis revealed cedrol-triggered release of cytochrome c, activation of caspase-9, and subsequent caspase-3 activation, initiating a caspase cascade reaction. This groundbreaking study establishes cedrol as the first compound to induce apoptosis in P. noxius while inhibiting its growth through oxidative stress, an increase in mitochondrial membrane permeability, and activation of the mitochondrial pathway. The findings offer compelling evidence for cedrol's potential as an effective antifungal agent against the destructive brown root disease caused by P. noxius.

Savinin Triggers Programmed Cell Death of Ray Parenchyma Cells in Heartwood Formation of Taiwania cryptomerioides Hayata.

The purpose of this study was to investigate the relationship between lignan biosynthesis and programmed cell death (PCD) of ray parenchyma cells during the heartwood formation of Taiwania (Taiwania cryptomerioides Hayata). Since the PCD of ray parenchyma cells and the synthesis of lignans are the two main processes involved in the formation of heartwood, both of which need to be completed through gene regulation. Based on the results of genomics and bioinformatics analysis, that the PCD of tracheids are induced by genotoxic, and the PCD of ray parenchyma cells is induced by biological factors, such as fungi, bacteria, and viruses, which could induce oxidative stress. According to the results of time-of-flight secondary ion mass spectrometry (ToF-SIMS) analysis, lignans are produced in ray parenchyma cells, and the accumulation of savinin and its downstream lignans might be the cause of PCD in ray parenchyma cells. An in vitro experiment further confirmed that the accumulation of savinin could cause protoplasts of Taiwania's xylem to produce taiwanin A, which is the marker of heartwood formation in Taiwania. Resulting in an increase in reactive oxygen species (ROS) content, which could induce oxidative stress in ray parenchyma cells and potentially lead to PCD. Based on these findings, we conclude that accumulation of savinin could be induced PCD of ray parenchyma cells in heartwood formation in Taiwania.

Two new diprenylated flavanones from Derris laxiflora Benth.

Phytochemical reinvestigation on the whole plants of Derris laxiflora Benth. afforded two new diprenylated flavanones, derriflavanones B and C (1-2), together with thirty-two known compounds, including sixteen flavonoids (3-18), eleven aromatic compounds (19-29), and five chlorophylls (30-34). All known compounds were first isolated from this plant. The structures of these compounds were determined by analysis of the NMR spectroscopy, mass data, IR spectra, UV spectra, optical rotation and by comparison with literature data.

New Anti-inflammatory Flavonol Glycosides from Lindera akoensis Hayata.

Inflammation is related to many diseases. Lindera akoensis Hayata was often used in folktherapy in Taiwan for inflammation. In this study, three new flavonol acyl glycosides, namelykaempferol-3-O--D-4",6"-di-(E)-p-coumaroylglucoside (1), 3"-(E)-p-coumaroylafzelin (2) and 40-Omethyl-2",4"-di-(E)-p-coumaroylquercitrin (3), and three components, 3-dodecyl-4-hydroxy-5-methyldihydrofuran-2-one (4), 2-acetoxyclovan-9-ol (5), (1,4,6)-trihydroxyeudesmane(6) that were isolated from the natural product for the first time were obtained along with 25 knowncompounds from L. akoensis. Their structures were determined by comprehensive spectroscopicanalyses (1D and 2D NMR, EI-, ESI- and HRESI-MS). The ability of 1 to decrease the LPS-stimulatedproduction of nitrite in RAW264.7 cell was evaluated, showing an IC50 value of 36.3 ± 3.2 µM.This result supports the value of L. akoensis as a traditional medicine resource.

Nonvolatile Taste Components and Antioxidant Properties of Fruiting Body and Mycelium with High Ergothioneine Content from the Culinary-Medicinal Golden Oyster Mushroom Pleurotus citrinopileatus (Agaricomycetes).

Pleurotus citrinopileatus mycelium was prepared with high ergothioneine (Hi-Ergo) content and its proximate composition, nonvolatile taste components, and antioxidant properties were studied. The ergothioneine contents of fruiting bodies and Hi-Ergo and regular mycelia were 3.89, 14.57, and 0.37 mg/g dry weight, respectively. Hi-Ergo mycelium contained more dietary fiber, soluble polysaccharides, and ash but less carbohydrates, reducing sugar, fiber, and fat than regular mycelium. However, Hi-Ergo mycelium contained the smallest amounts of total sugars and polyols (47.43 mg/g dry weight). In addition, Hi-Ergo mycelium showed the most intense umami taste. On the basis of the half-maximal effective concentration values obtained, the 70% ethanolic extract from Hi-Ergo mycelium showed the most effective antioxidant activity, reducing power, and scavenging ability, whereas the fruiting body showed the most effective antioxidant activity, chelating ability, and Trolox-equivalent antioxidant capacity. Overall, Hi-Ergo mycelium could be beneficially used as a food-flavoring material or as a nutritional supplement.

Quantification of Water-Soluble Metabolites in Medicinal Mushrooms Using Proton NMR Spectroscopy.

The water-soluble metabolites in 5 mushrooms were identified and quantified using proton nuclear magnetic resonance (NMR) spectroscopy and software for targeted metabolite detection and quantification. In total, 35 compounds were found in Agaricus brasiliensis, 25 in Taiwanofungus camphoratus, 23 in Ganoderma lucidum (Taiwan) and Lentinus edodes, and 16 in G. lucidum (China). Total amounts of all identified metabolites in A. brasiliensis, T. camphoratus, G. lucidum, G. lucidum (China), and L. edodes were 149,950.51, 12,834.18, 9,549.09, 2,788.41, and 111,726.51 mg/kg dry weight, respectively. These metabolites were categorized into 4 groups: free amino acids and derivatives, carbohydrates, carboxylic acids, and nucleosides. Carbohydrates were the most abundant metabolites among all 4 groups, with mannitol having the highest concentration among all analyzed metabolites (848-94,104 mg/kg dry weight). Principal components analysis (PCA) showed obvious distinction among the metabolites of the 5 different kinds of mushrooms analyzed in this study. Thus PCA could provide an optional analytical way of identifying and recognizing the compositions of flavor products. Furthermore, the results of this study demonstrate that NMRbased metabolomics is a powerful tool for differentiating between various medicinal mushrooms.

Induction of IL-25 secretion from tumour-associated fibroblasts suppresses mammary tumour metastasis.

Tumour-associated fibroblasts (TAFs), as a functionally supportive microenvironment, play an essential role in tumour progression. Here we investigate the role of IL-25, an endogenous anticancer factor secreted from TAFs, in suppression of mouse 4T1 mammary tumour metastasis. We show that a synthetic dihydrobenzofuran lignan (Q2-3), the dimerization product of plant caffeic acid methyl ester, suppresses 4T1 metastasis by increasing fibroblastic IL-25 activity. The secretion of IL-25 from treated human or mouse fibroblasts is enhanced in vitro, and this activity confers a strong suppressive effect on growth activity of test carcinoma cells. Subsequent in vivo experiments showed that the anti-metastatic effects of Q2-3 on 4T1 and human MDA-MD-231 tumour cells are additive when employed in combination with the clinically used drug, docetaxel. Altogether, our findings reveal that the release of IL-25 from TAFs may serve as a check point for control of mammary tumour metastasis and that phytochemical Q2-3 can efficiently promote such anticancer activities.

Pterocarpans from Derris laxiflora.

Six compounds were isolated from Derris laxiflora Benth., including two new pterocarpans, 7,6'-dihydroxy-3'-methoxypterocarpan (1) and derrispisatin (2), as well as four known ones, lespedezol D, (3), secundiflorol 1 (4), 6a-hydroxymaackiain (5) and pisatin (6). The structures of these compounds were determined by analysis of their spectroscopic data.

Metabolite Profiling and Comparison of Bioactivity in Antrodia cinnamomea and Antrodia salmonea Fruiting Bodies.

Antrodia cinnamomea is a precious edible mushroom endemic to Taiwan that has been claimed to have significant health promotion activities. Antrodia salmonea is a new species of the genus Antrodia. In this study, we compared the metabolites and bioactivity of A. cinnamomea and A. salmonea fruiting bodies. The volatiles of A. cinnamomea and A. salmonea were characterized and 3,4,5-trimethoxybenzaldehyde was found to be the most abundant compound in A. cinnamomea; the other abundant compounds were δ-guaiene, isolongifolene, 1-octen-3-ol, 4-terpinenol, α-guaiene, and p-cymene. In A. salmonea, the main volatiles were α-cedrene, 1-octen-3-ol, D-limonene, cadinadiene, germacrene D, isolongifolene, and α-muurolene. Furthermore, five ergostane-type triterpenoids and two lanostane-type triterpenoids were selected as index compounds characterizing A. cinnamomea and A. salmonea extracts. The content of each compound varied between the two species. (R,S)-antcin B was the most abundant compound in A. cinnamomea fruiting bodies (75.18 ± 0.11 µg/mg). However, (R,S)-antcin C (184.85 ± 0.96 µg/mg) was the major triterpenoid in the A. salmonea fruiting body. Furthermore, two compounds, antcin M and methyl antcinate K, were only present in the A. salmonea fingerprint; therefore, antcin M and methyl antcinate K may be important for distinguishing between A. cinnamomea and A. salmonea fruiting bodies. Finally, examination of anti-inflammation activity and cytotoxicity showed that A. salmonea had more anti-inflammatory activity than A. cinnamomea; however, A. salmonea was more cytotoxic than A. cinnamomea. In conclusion, the composition and bioactivity of the fruiting bodies of A. cinnamomea and A. salmonea varies. Therefore, it is recommended that further toxicological evaluation and investigation of the biological activity of A. salmonea is carried out to ensure its safe and efficacious use as an alternative to A. cinnamomea.

Submerged Cultivation of Mycelium with High Ergothioneine Content from the Culinary-Medicinal Golden Oyster Mushroom, Pleurotus citrinopileatus (Higher Basidiomycetes).

The optimization of submerged culture of the culinary-medicinal golden oyster mushroom, Pleurotus citrinopileatus, was studied using a one-factor-at-a-time, two-stage stimulation and central composite rotatable design to produce mycelia with high ergothioneine content. The optimal culture conditions for mycelia harvested at day 22 were a temperature of 25°C, an inoculation ratio of 5%, 2% glucose, 0.5% yeast extract, and adjustment of the initial pH value to 10. The biomass and ergothioneine content were 8.28 g/L and 10.65 mg/g dry weight (dw), respectively. The addition of an amino acid precursor increased the ergothioneine content of mycelia; cysteine was the most effective. In addition, the results obtained from central composite rotatable design showed that the recommended combination for cysteine, histidine, and methionine was 8, 4, and 0.5 mmol/L, respectively. The predicted ergothioneine content was 13.90 mg/g dw, whereas the experimental maximal ergothioneine content was 14.57 mg/g dw. With the addition of complex precursors and under optimal culture conditions, mycelia harvested at days 16-20 had higher ergothioneine content. Accordingly, the information obtained could be used to produce mycelia with high ergothioneine content.

Effect of Hinoki and Meniki Essential Oils on Human Autonomic Nervous System Activity and Mood States.

Meniki (Chamecyparis formosensis) and Hinoki (C. obtusa) are precious conifers with excellent wood properties and distinctive fragrances that make these species popular in Taiwan for construction, interiors and furniture. In the present study, the compositions of essential oils prepared from Meniki and Hinoki were analyzed by gas chromatography-mass spectrometry (GC/MS). Thirty-six compounds were identified from the wood essential oil of Meniki, including Δ-cadinene, γ-cadinene, Δ-cadinol, α-muurolene, calamenene, linalyl acetate and myrtenol; 29 compounds were identified from Hinoki, including α-terpineol, α-pinene, Δ-cadinene, borneol, terpinolene, and limonene. Next, we examined the effect of Meniki and Hinoki essential oils on human autonomic nervous system activity. Sixteen healthy adults received Meniki or Hinoki by inhalation for 5 min, and the physiological and psychological effects were examined. After inhaling Meniki essential oil, participant's systolic blood pressure and heart rate (HR) were decreased, and diastolic blood pressure increased. In addition, sympathetic nervous activity (SNS) was significantly decreased, and parasympathetic activity (PSNS) was significantly increased. On the other hand, after inhaling Hinoki essential oil, systolic blood pressure, heart rate and PSNS were decreased, whereas SNA was increased. Indeed, both Meniki and Hinoki essential oils increased heart rate variability (HRV) in tested adults. Furthermore, in the Profile of Mood States (POMS) test, both Meniki and Hinoki wood essential oils stimulated a pleasant mood status. Our results strongly suggest that Meniki and Hinoki essential oils could be suitable agents for the development of regulators of sympathetic nervous system dysfunctions.

Naturally occurring anthraquinones: chemistry and therapeutic potential in autoimmune diabetes.

Anthraquinones are a class of aromatic compounds with a 9,10-dioxoanthracene core. So far, 79 naturally occurring anthraquinones have been identified which include emodin, physcion, cascarin, catenarin, and rhein. A large body of literature has demonstrated that the naturally occurring anthraquinones possess a broad spectrum of bioactivities, such as cathartic, anticancer, anti-inflammatory, antimicrobial, diuretic, vasorelaxing, and phytoestrogen activities, suggesting their possible clinical application in many diseases. Despite the advances that have been made in understanding the chemistry and biology of the anthraquinones in recent years, research into their mechanisms of action and therapeutic potential in autoimmune disorders is still at an early stage. In this paper, we briefly introduce the etiology of autoimmune diabetes, an autoimmune disorder that affects as many as 10 million worldwide, and the role of chemotaxis in autoimmune diabetes. We then outline the chemical structure and biological properties of the naturally occurring anthraquinones and their derivatives with an emphasis on recent findings about their immune regulation. We discuss the structure and activity relationship, mode of action, and therapeutic potential of the anthraquinones in autoimmune diabetes, including a new strategy for the use of the anthraquinones in autoimmune diabetes.

Anti-inflammatory and anti-oxidative activities of polyacetylene from Dendropanax dentiger.

Dendropanax dentiger has been used as a folk medicine since ancient times. In our current study, we observed that D. dentiger exhibited a significant anti-inflammatory activity, which could efficiently inhibit nitric oxide (NO) production in the lipopolysaccharide (LPS)-induced macrophage inflammation assay. (9Z,16S)-16-Hydroxy-9,17-octadecadiene-12,14-diynoic acid (HODA) was isolated from the leaves of D. dentiger following a bioactivity guided fractionation protocol. Our data indicated that HODA significantly inhibited the NO production in LPS-induced RAW 264.7 murine macrophage cells (IC50 = 4.28 µM). Consistent with these observations, the mRNA and protein expression levels of iNOS were also inhibited by HODA in a dose-dependent manner. HODA also reduced the translocation of NF-κB into nuclear fractions. Meanwhile, HODA enhanced Nrf-2 activation and its downstream antioxidant gene HO-1. We concluded that HODA possessed significant anti-inflammatory and anti-oxidative activity; the compound may have a potential for development as a chemoprevention agent.

A plant kavalactone desmethoxyyangonin prevents inflammation and fulminant hepatitis in mice.

Alpinia pricei Hayata is a Formosan plant which has been popularly used as nutraceutical or folk medicine for inflammation and various disorders. An active compound of the plant rhizomes, desmethoxyyangonin (DMY), was identified in this study for its novel effect against endotoxin lipopolysaccharide (LPS)-stimulated inflammation in murine macrophages and LPS/D-galactosamine (LPS/D-GalN)-induced fulminant hepatitis in mice. DMY was observed to significantly inhibit proliferation and activation of T cells ex vivo and the activity of several pro-inflammatory mediators in vitro. DMY also protected LPS/D-GalN-induced acute hepatic damages in mice through inhibiting aminotransferases activities and infiltrations of inflammatory macrophages, neutrophils and pathogenic T cells into the liver tissues. In addition, pretreatment with DMY significantly improved the survival rate of LPS/D-GalN-treated mice to 90% (9/10), compared to LPS/D-GalN-treated group (40%, 4/10). UPLC/MS platform-based comparative metabolomics approach was used to explore the serum metabolic profile in fulminant hepatic failure (FHF) mice with or without the DMY pretreatment. The results showed that LPS/D-GalN-induced hepatic damage is likely through perturbing amino acid metabolism, which leads to decreased pyruvate formation via catalysis of aminotransferases, and DMY treatment can prevent to a certain degree of these alterations in metabolic network in mouse caused by LPS/D-GalN. Mechanistic investigation demonstrated that DMY protects LPS or LPS/D-GalN-induced damages in cell or liver tissues mainly through de-regulating IKK/NFκB and Jak2/STAT3 signaling pathways. This report provides evidence-based knowledge to support the rationale for the use of A. pricei root extract in anti-inflammation and also its new function as hepatoprotetive agent against fulminant hepatitis.

Establishment of the metabolite profile for an Antrodia cinnamomea health food product and investigation of its chemoprevention activity.

Antrodia cinnamomea is an edible fungus endemic to Taiwan that has been attributed with health promotion benefits. An A. cinnamomea mycelium health food product, which was produced by solid-state culture, was selected as the target for investigation in this study. Fourteen representative metabolites of A. cinnamomea mycelium (EMAC) were selected as index compounds to establish the metabolite profile for evaluation of EMAC product quality. It was also demonstrated that EMAC administration significantly reduced liver inflammation and serum oxidative stress in vivo. 4-Acetylantroquinonol B obtained by a bioactivity-guided fractionation from EMAC was able to not only inhibit LPS-induced nitric oxide formation in macrophages but also protect against ethanol-induced oxidative stress in liver cells. The results suggest this A. cinnamomea product might be a potent antioxidative and anti-inflammatory supplement for chemoprevention.

The effect of the aerial part of Lindera akoensis on lipopolysaccharides (LPS)-induced nitric oxide production in RAW264.7 cells.

Four new secondary metabolites, 3α-((E)-Dodec-1-enyl)-4ß-hydroxy-5ß-methyldihydrofuran-2-one (1), linderinol (6), 4'-O-methylkaempferol 3-O-α-L-(4''-E-p-coumaroyl)rhamnoside (11) and kaempferol 3-O-α-L-(4''-Z-p-coumaroyl)rhamnoside (12) with eleven known compounds-3-epilistenolide D1 (2), 3-epilistenolide D2 (3), (3Z,4α,5ß)-3-(dodec-11-ynylidene)-4-hydroxy-5-methylbutanolide (4), (3E,4ß,5ß)-3-(dodec-11-ynylidene)-4-hydroxy-5-methylbutanolide (5), matairesinol (7), syringaresinol (8), (+)-pinoresinol (9), salicifoliol (10), 4''-p-coumaroylafzelin (13), catechin (14) and epicatechin (15)-were first isolated from the aerial part of Lindera akoensis. Their structures were determined by detailed analysis of 1D- and 2D-NMR spectroscopic data. All of the compounds isolated from Lindera akoensis showed that in vitro anti-inflammatory activity decreases the LPS-stimulated production of nitric oxide (NO) in RAW 264.7 cell, with IC50 values of 4.1-413.8 µM.

Anti-inflammatory lanostanoids and lactone derivatives from Antrodia camphorata.

Four new lanostanoids, ethyl lucidenate A (1), ethyl lucidenate F (2), 15-O-acetylganolucidate A (3), and 3,11,15,23-tetraoxo-27ξ-lanosta-8,16-dien-26-oic acid (4), and two new lactone derivatives, 5-hydroxy-5-(methoxymethyl)-4-methylfuran-2(5H)-one (5) and 3-(4-methoxy-2-oxo-2H-pyran-6-yl)propanoic acid (6), together with four known compounds, 11α-hydroxy-3,7-dioxolanost-8,24(E)-dien-26- oic acid (7), 3,7,11-trioxo-5α-lanosta-8,24(E)-dien-26-oic acid (8), methyl 3,7,11,12,15,23-hexaoxo-5α-lanost-8-en-26-oate (9), and ethyl 3,7,11,12,15,23-hexaoxo-5α-lanost-8-en-26-oate (10), were characterized from Antrodia camphorata. The structures of these new compounds were determined by analysis of their spectroscopic data, including 1D and 2D NMR experiments. Ten components were evaluated for anti-inflammatory activity by examining their effect on LPS-iNOS-dependent NO production in murine macrophage (RAW 264.7) cells. Among them, compounds 1, 3, 7, 8, 9, and 10 significantly suppressed the NO concentration in LPS-treated RAW 264.7 cells with IC50 values ≤ 10 µM.

Antcin C from Antrodia cinnamomea Protects Liver Cells Against Free Radical-Induced Oxidative Stress and Apoptosis In Vitro and In Vivo through Nrf2-Dependent Mechanism.

In this study, we investigated the cytoprotective effects of antcin C, a steroid-like compound isolated from Antrodia cinnamaomea against AAPH-induced oxidative stress and apoptosis in human hepatic HepG2 cells. Pretreatment with antcin C significantly protects hepatic cells from AAPH-induced cell death through the inhibition of ROS generation. Furthermore, AAPH-induced lipid peroxidation, ALT/AST secretion and GSH depletion was significantly inhibited by antcin C. The antioxidant potential of antcin C was correlated with induction of antioxidant genes including, HO-1, NQO-1, γ -GCLC, and SOD via transcriptional activation of Nrf2. The Nrf2 activation by antcin C is mediated by JNK1/2 and PI3K activation, whereas pharmacologic inhibition of JNK1/2 and PI3K abolished antcin C-induced Nrf2 activity. In addition, AAPH-induced apoptosis was significantly inhibited by antcin C through the down-regulation of pro-apoptotic factors including, Bax, cytochrome c, capase 9, -4, -12, -3, and PARP. In vivo studies also show that antcin C significantly protected mice liver from AAPH-induced hepatic injury as evidenced by reduction in hepatic enzymes in circulation. Further, immunocytochemistry analyses showed that antcin C significantly increased HO-1 and Nrf2 expression in mice liver tissues. These results strongly suggest that antcin C could protect liver cells from oxidative stress and cell death via Nrf2/ARE activation.

Lanostane triterpenoids and sterols from Antrodia camphorata.

Four lanostane triterpenes, 3,7,11-trioxo-5α-lanosta-8,24(E)-dien-26-oic acid, methyl 11α-3,7-dioxo-5α-lanosta-8,24(E)-dien-26-oate, methyl 3,7,11,12,15,23-hexaoxo-5α-lanost-8-en-26-oate, and ethyl 3,7,11,12,15,23-hexaoxo-5α-lanost-8-en-26-oate, two sterols, (14α,22E)-14-hydroxyergosta-7,22-diene-3,6-dione and a steroid named as camphosterol A were isolated from a mixture of fruiting bodies and mycelia of solid cultures of Antrodia camphorata. The ¹H and ¹³C NMR spectra of all compounds were fully assigned using a combination of 2D NMR experiments, including COSY, HMQC, HMBC and NOESY sequences. Six compounds were evaluated for cytotoxicity against several human tumor cell lines, all of which has moderate activity.