As part of our continuous effort to find potential anti-inflammatory agents from endophytic fungi, a Fusarium solani strain, isolated from the plant Aponogeton undulatus Roxb., was investigated. Cerevisterol (CRVS) was identified from endophytic fungi, a Fusarium solani strain, and moreover exhibited anti-inflammatory activity. However, the underlying mode of action remains poorly understood. The aim of this study is to reveal the potential mechanisms of CRVS against inflammation on a molecular level in LPS-activated RAW 264.7 peritoneal macrophage cells. CRVS was isolated from F. solani and characterized based on spectral data analysis. The MTT assay was performed to measure cell viability in CRVS-treated macrophages. Anti-inflammatory activity was assessed by measurement of nitric oxide (NO) and prostaglandin E2 (PGE2) levels, as well as the production of various cytokines, such as tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β), and -6 (IL-6) in LPS-stimulated macrophages. RT-PCR and immunoblotting analyses were done to examine the expression of various inflammatory response genes. A reporter gene assay was conducted to measure the level of nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) and activator protein-1 (AP-1) transactivation. CRVS suppresses the LPS-induced production of NO and PGE2, which is a plausible mechanism for this effect is by reducing the expression of iNOS and COX-2. CRVS also decreases the expression of pro-inflammatory cytokines, such as TNF-α, IL-6, and IL-1β. CRVS halted the nuclear translocation of NF-κB by blocking the phosphorylation of inhibitory protein κBα (IκBα) and suppressing NF-κB transactivation. The mitogen-activated protein kinases (MAPK) signaling pathways are also suppressed. CRVS treatment also inhibited the transactivation of AP-1 and the phosphorylation of c-Fos. Furthermore, CRVS could induce the nuclear translocation of nuclear factor erythroid 2-related factor 2 (Nrf2) by down-regulating Kelch-like ECH-associated protein 1 (Keap-1) and up-regulating hemeoxygenases-1 (HO-1) expression. The results suggest that CRVS acts as a natural agent for treating inflammatory diseases by targeting an MAPK, NF-κB, AP-1, and Nrf2-mediated HO-1 signaling cascade.

Cladosporium species are endophytic fungi that grow on organic matter and are considered food contaminants. The anti-microbial and anti-tumor naphthoquinones fusarubin (FUS) and anhydrofusarubin (AFU) were isolated using column chromatography from a Cladosporium species residing inside Rauwolfia leaves. The impact of FUS and AFU on cell growth was assessed in acute myeloid leukemia (OCI-AML3) and other hematologic tumor cell lines (HL-60, U937, and Jurkat). Treatment with FUS or AFU reduced the number of OCI-AML3 cells as evaluated by a hemocytometer. Flow cytometry analyses showed that this effect was accompanied by diverse impairments in cell cycle progression. Specifically, FUS (20 or 10 μg/mL significantly decreased the percentage of cells in S phase and increased the percentage of cells in G2/M phase, whereas AFU increased the percentage of cells in G0/G1 phase (50 and 25 μg/mL) and decreased the percentage of cells in S (50 μg/mL) and G2/M (50 and 25 μg/mL) phases. Both substances significantly increased apoptosis at higher concentrations. The effects of FUS were more potent than those of AFU, with FUS up-regulating p21 expression in a p53-dependent manner, as detected by Western blot analyses, likely the consequence of decreased ERK phosphorylation and increased p38 expression (both of which increase p21 stability). FUS also decreased Akt phosphorylation and resulted in increased Fas ligand production and caspase-8/3-dependent apoptosis. These results suggest that FUS and AFU inhibit proliferation and increase apoptosis in cell lines derived from hematological cancers.

Three novel flavonoid glycosides, 5,6-dimethyoxy-3',4''-dioxymethylene-7-O-(6''-beta-D-glucopyranosyl-beta-D-glucopyranosyl) flavanone (1), 5,4'-dihydroxy-3-O-alpha-L-rhamnopyranosyl-6-C-glucopyranosyl-7-O-(6''-para-coumaroyl-beta-D-glucopyranosyl) flavone (2) and 5,4'-dihydroxy-3-O-(2'''''-beta-glucopyranosyl-alpha-L-rhamnopyranosyl)-6-C-glucopyranosyl-7-O-(6''-para-coumaroyl-beta-D-glucopyranosyl) flavone (3) were isolated from the 1-butanol soluble fraction of the bulbs of the plant Urginea indica (Indian squill). The structures of the compounds were elucidated on the basis of spectral analysis, including homo- and heteronuclear correlation NMR experiments (COSY, NOESY, HSQC and HMBC) and mass spectra.

Xylocarpus granatum, a common mangrove plant is traditionally used for the treatment of diarrhoea, cholera, fever, dyslipidemia, inflammation, etc. The present study was carried out to evaluate the antioxidant and anti-inflammatory potential of the ethanolic extract of X. Granatum by various in vitro methods such as 1,1-diphenyl-2-picryl-hydrazil (DPPH) free radical scavenging assay,  reducing power assay, ferric reducing antioxidant power (FRAP) and β-carotene bleaching inhibition assay. Total phenolic and flavonoid content were determined. Anti-inflammatory activity was evaluated by in vitro human RBC membrane stabilizing assay and in vivo mice paw edema test. Ethanolic leaf extract (S2) of X. granatum showed significant scavenging effect on DPPH scavenging with a value of IC50 165.95µg/ml. In addition, it showed significant reducing potential with a value of 59.04 mM of ferrous equivalent per ml in FRAP assay and in reducing power assay the EC50 value was determined as 241.61μg/ml . The ethanolic leaf extracts exhibited 72.3% β-carotene bleaching inhibition. The total phenolic and flavonoid content of the extract were 66μg/ml gallic acid equivalent and 47.66μg/ml quercetin equivalent per gram of dry extract, respectively. The extract also exhibited 52.63% and 51.05% protection of RBC membrane in hypotonicity and heat induced lysis inhibition, respectively. Significant reduction of mice paw edema (36.34% in 20 μg/kg bw concentration) was observed in the extract. The results revealed that the leaf extract of X. granatum possesses strong antioxidant and anti-inflammatory potential. J Bangladesh Agril Univ 17(4): 466–475, 2019

Three steroids, namely 24-ethyl-5α-cholestan-3-one (1), 5α-stigmast-22-en-3-one (2), stigmast-5, 22-dien-3-one (3) have been isolated from N. stellata. The phytochemical and antimicrobial as well as cytotoxic activities of Nymphaea stellata were investigated in this study. Crude extracts of N. stellata and various column fractions exhibited poor antimicrobial activity against a wide range of Gram-positive and Gram-negative bacteria and fungi. The crude extract and the fractions showed significant cytotoxic effect when subjected to brine shrimp lethality bioassay.

Background Endophytic fungi play a vital role in plant defense system by secreting various antimicrobial agents. To evaluate the antimicrobial activity of the endophytic fungi of the mangrove plant Heritiera fomes (Buch. -Ham), plants were collected from the Sundarbans, Bangladesh. The endophytic fungi were subsequently isolated and identified. Results Fifty-five endophytic fungi were isolated from the leaves, root, and bark of H. fomes (Buch. -Ham). Genomic DNA was extracted for PCR (polymerase chain reaction) by specific primers to multiply ITS region and sequences were determined. Nucleotide sequence data were submitted to the Genbank and accession number for each fungal strain was obtained. Antimicrobial activity of the ethyl acetate (EtOAc) and methanolic extracts of eleven species from both fermentation and mycelium, respectively, were analyzed by microtiter plate-based antimicrobial assay incorporating resazurin as an indicator of cell growth against two Gram-positive bacteria namely Staphylococcus aureus NCTC 12981 and Micrococcus luteus NCTC 7508, two Gram-negative bacteria namely Escherichia coli NCTC 12241 and Pseudomonas aeruginosa NCTC 12903, and a fungus Candida albicans ATCC 90028. All the endophytic fungal extracts exhibited antimicrobial activities against more than one-tested pathogenic microbial strains. Overall, methanolic extracts showed greater activity than EtOAc extracts. Pseudopestalotiopsis camelliae-sinensis, Pestalotiopsis microspora, and Penicillium copticola were the most active endophytic fungal strains and exhibited strong inhibitory activity against the microorganisms under investigation and their MIC values ranged from 0.0024 to 5.0 mg/mL. Methanolic extracts of both P. camelliae-sinensis and P. microspora showed the highest antibacterial activity (MIC value of 0.0024 mg/mL) against P. aeruginosa NCTC 12903. Conclusion This study showed that the isolated and identified endophytic fungi from H. fomes (Buch. -Ham) could be potential sources of antimicrobial agents.

Background:Ravenia spectabilis is a medium tall shrub found widespread in South America. It also found in India, Pakistan, Bangladesh etc. Few alkaloid and steroid compounds were reported from the plant previously.Materials and Methods:Methanol extract from the stems of Ravenia spectabilis were partitioned into n-hexane, carbon tetrachloride, chloroform and aqueous soluble fractions, respectively. The crude methanol extract, carbon tetrachloride fraction and chloroform fraction were fractionated by column chromatography of Silica gel and Sephadex LH-20 for isolation and purification of compounds. The structures of the isolated compounds were determined by extensive NMR spectral analysis, including 2D NMR, mass spectroscopy etc.Results:Ten compounds, γ-fagarine (1), ravenoline (2), N-methyl atanine (3),2,3,3,5-tetramethyl-2,3,4,5- tetrahydrofurano [3,2-c] quinolin-4-one (4), arborinine (5), 3-geranyl indole (6), atanine (7), steroids sitosta-4-en- 3-one (8), stigmasterol (9) and 3-methoxy-4-hydroxy cinnamic acid (10) were isolated from the stems of Ravenia spectabilis.Conclusion:Compounds N-methyl atanine (3), 2,3,3,5-tetramethyl-2,3,4,5-tetrahydrofurano [3,2-c] quinolin-4-one (4), 3-geranyl indole (6), sitosta-4-en-3-one (8) and 3-methoxy-4-hydroxy cinnamic acid (10) were isolated from this plant for the first time. 3-geranyl indole (6) was also isolated second time from natural sources.

The crude methanolic extract of the root bark of Pongamia pinnata was taken into consideration to isolate secondary metabolites. A total six known natural compounds were separated and purified by various chromatographic techniques and five isolates were identified as flavonoid derivatives such as pongachromene (1), kanugin (2), karanjin (3), demethoxykanugin (4), dimethoxypongapine (5) and the other is a triterpenoid (6). The pure compounds as well as petroleum ether, dichloromethane and ethyl acetate soluble fractions of crude methanolic extract were evaluated for bioactivities using established methods. In vitro antioxidant activity was studied by DPPH radical scavenging method using butylated hydroxyl anisole as standard. Among the pure compounds, kanugin and pongachromene showed significant antioxidant activity with the IC50 values of 27.20 ± 0.39 μg/mL and 43.53 ± 0.63 μg/ml, respectively as compared to the standard (23.87 ± 0.09 μg/ml), whereas karanjin, demethoxykanugin and dimethoxypongapine demonstrated moderate antioxidant activity. Mild thrombolytic activity was observed by different fractions with clot lysis ranging from 18.49 to 29.35% as compared to standard streptokinase (79.12%). The different solvent fractions and pure isolates showed very mild antimicrobial activity with zone of inhibition of 7.5 - 10.0 mm against the tested microorganisms using azithromycin and ketoconazole as standards. In the brine shrimp lethality bioassay, the dichloromethane, ethyl acetate, and methanol soluble fractions revealed significant lethality with LC50 values of 0.67 ± 0.05, 0.61 ± 0.13 and 0.56 ± 0.10 μg/ml, respectively as compared to standard tamoxifen (LC50 value 0.34 ± 0.09 μg/ml). Dhaka Univ. J. Pharm. Sci. 19(1): 1-8, 2020 (June)