Natural Polyketides Isolated from the Endophytic Fungus Phomopsis sp. CAM212 with a Semisynthetic Derivative Downregulating the ERK/IκBα Signaling Pathways.

Three previously undescribed natural products, phomopsinin A - C (1:  - 3: ), together with three known compounds, namely, cis-hydroxymellein (4: ), phomoxanthone A (5: ) and cytochalasin L-696,474 (6: ), were isolated from the solid culture of Phomopsis sp. CAM212, an endophytic fungus obtained from Garcinia xanthochymus. Their structures were determined on the basis of spectroscopic data, including IR, NMR, and MS. The absolute configurations of 1: and 2: were assigned by comparing their experimental and calculated ECD spectra. Acetylation of compound 1: yielded 1A: , a new natural product derivative that was tested together with other isolated compounds on lipopolysaccharide-stimulated RAW 264.7 cells. Cytochalasin L-696,474 (6: ) was found to significantly inhibit nitric oxide production, but was highly cytotoxic to the treated cells, whereas compound 1: slightly inhibited nitric oxide production, which was not significantly different compared to lipopolysaccharide-treated cells. Remarkably, the acetylated derivative of 1: , compound 1A: , significantly inhibited nitric oxide production with an IC50 value of 14.8 µM and no cytotoxic effect on treated cells, thereby showing the importance of the acetyl group in the anti-inflammatory activity of 1A: . The study of the mechanism of action revealed that 1A: decreases the expression of inducible nitric oxide synthase, cyclooxygenase 2, and proinflammatory cytokine IL-6 without an effect on IL-1ß expression. Moreover, it was found that 1A: exerts its anti-inflammatory activity in lipopolysaccharide-stimulated RAW 264.7 macrophage cells by downregulating the activation of ERK1/2 and by preventing the translocation of nuclear factor κB. Thus, derivatives of phomopsinin A (1: ), such as compound 1A: , could provide new anti-inflammatory leads.

Lambertellin from Pycnoporus sanguineus MUCL 51321 and its anti-inflammatory effect via modulation of MAPK and NF-κB signaling pathways.

Lambertellin (1) and ergosta-5,7,22-trien-3-ol (2) were isolated from the solid rice fermentation of the plant pathogenic fungus Pycnoporus sanguineus MUCL 51321. Their structures were elucidated using comprehensive spectroscopic methods. The isolated compounds were tested on lipopolysaccharide (LPS)-stimulated RAW 264.7 macrophage cells. Lambertellin (1) exhibited promising inhibitory activity against nitric oxide (NO) production with IC50 value of 3.19 µM, and it significantly inhibited the expression of inducible NO synthase (iNOS) and cyclooxygenase 2 (COX-2). Lambertellin (1) also decreased the expression of pro-inflammatory cytokines IL-6 and IL-1ß. The study of the mechanistic pathways revealed that lambertellin (1) exerts its anti-inflammatory effect in LPS-stimulated RAW 264.7 macrophage cells by modulating the activation of the mitogen activated protein kinase (MAPK) and nuclear factor κB (NF-κB) signaling pathways. Therefore, lambertellin (1) could be a promising lead compound for the development of new anti-inflammatory drugs.

Antibacterial and cytotoxic cytochalasins from the endophytic fungus Phomopsis sp. harbored in Garcinia kola (Heckel) nut.

BACKGROUND: The continuous emergence of multidrug-resistant (MDR) bacteria drastically reduced the efficacy of our antibiotic armory and consequently, increased the frequency of therapeutic failure. The search for bioactive constituents from endophytic fungi against MDR bacteria became a necessity for alternative and promising strategies, and for the development of novel therapeutic solutions. We report here the isolation and structure elucidation of antibacterial and cytotoxic compounds from Phomopsis sp., an endophytic fungus associated with Garcinia kola nuts. METHODS: The fungus Phomopsis sp. was isolated from the nut of Garcinia kola. The crude extract was prepared from mycelium of Phomopsis sp. by maceration in ethyl acetate and sequentially fractionated by column chromatography. The structures of isolated compounds were elucidated on the basis of spectral studies and comparison with published data. The isolated compounds were evaluated for their antibacterial and anticancer properties by broth microdilution and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide methods respectively. The samples were also tested spectrophotometrically for their hemolytic properties against human red blood cells. RESULTS: The fractionation of the crude extract afforded three known cytochalasins including 18-metoxycytochalasin J (1), cytochalasins H (2) and J (3) together with alternariol (4). The cytochalasin compounds showed different degrees of antibacterial activities against the tested bacterial pathogens. Shigella flexneri was the most sensitive microorganism while Vibrio cholerae SG24 and Vibrio cholerae PC2 were the most resistant. Ampicillin did not show any antibacterial activity against Vibrio cholerae NB2, Vibrio cholerae PC2 and Shigella flexneri at concentrations up to 512 µg/mL, but interestingly, these multi-drug resistant bacterial strains were sensitive to the cytochalasin metabolites. These compounds also showed significant cytotoxic properties against human cancer cells (LC50 = 3.66-35.69 µg/mL) with low toxicity to normal non-cancer cells. CONCLUSION: The three cytochalasin compounds isolated from the Phomopsis sp. crude extract could be a clinically useful alternative for the treatment of cervical cancer and severe infections caused by MDR Shigella and Vibrio cholerae.

One new constituent from the stem bark of Chrysophyllum lacourtianum De Wild. (sapotaceae).

The phytochemical investigation of the methanol extract of the stem bark of Chrysophyllum lacourtianum led to the isolation and characterization of one new secondary metabolite, lacourtianal (1), together with eight known compounds. Compounds 2; 3; 5; 6; 7 and 9 were reported for the first time from Chrysophyllum genus. The structures of compounds 1-9 were elucidated on the basis of 1 D and 2 D NMR spectroscopic and mass spectrometric data as well as comparison with the literature. The antibacterial activity of the methanol extract, fractions and compounds 1-9 were evaluated against bacterial strains. The methanol extract exhibited moderate activity against Staphylococcus aureus NR4674 with MIC values of 500 µg/mL. The n-hexane fraction showed moderate activity against Staphylococcus aureus (ATCC 43300 and ATCC 25923) with MIC values of 125 µg/mL and ursolic acid (5) exhibited strong activity against Enterobacter aerogenes CPC and Escherichia coli ATCC 25322 with MIC values of 7.8 and 3.9 µg/mL respectively.

Evaluation of anti-quorum sensing and antibiofilm effects of secondary metabolites from Gambeya lacourtiana (De Wild) Aubr. & Pellegr against selected pathogens.

BACKGROUND: Microbial infections cause serious health problems especially with the rising antibiotic resistance which accounts for about 700,000 human deaths annually. Antibiotics which target bacterial death encounter microbial resistance with time, hence, there is an urgent need for the search of antimicrobial substances which target disruption of virulence factors such as biofilm and quorum sensing (QS) with selective pressure on the pathogens so as to avoid resistance. METHODS: Natural products are suitable leads for antimicrobial drugs that can inhibit bacterial biofilms and QS. Twenty compounds isolated from the medicinal plant Gambeya lacourtiana were evaluated for their antibiofilm and anti-quorum sensing effects against selected pathogenic bacteria. RESULTS: Most of the compounds inhibited violacein production in Chromobacterium violaceum CV12472 and the most active compound, Epicatechin had 100% inhibition at MIC (Minimal Inhibitory Concentration) and was the only compound to inhibit violacein production at MIC/8 with percentage inhibition of 17.2 ± 0.9%. Since the bacteria C. violaceum produces violacein while growing, the inhibition of the production of this pigment reflects the inhibition of signal production. Equally, some compounds inhibited violacein production by C. violaceum CV026 in the midst of an externally supplied acylhomoserine lactone, indicating that they disrupted signal molecule reception. Most of the compounds exhibited biofilm inhibition on Staphyloccocus aureus, Escherichia coli and Candida albicans and it was observed that the Gram-positive bacteria biofilm was most susceptible. The triterpenoids bearing carboxylic acid group, the ceramide and epicatechin were the most active compounds compared to others. CONCLUSION: Since some of the compounds disrupted QS mediated processes in bacteria, it indicates that this plant is a source of antibiotics drugs that can reduce microbial resistance.

A new sphingolipid and furanocoumarins with antimicrobial activity from Ficus exasperata.

From the methanol extract of the stem bark of Ficus exasperata, a new sphingolipid named Ficusamide, (2S,3S,4R,11E)-2-[(2',3'-dihydroxyhexacosanoylamino)]-11-octadecene-1,3,4-triol (1), along with three known furanocoumarins, (S)-(-) oxypeucedanin hydrate (2), (R)-(+) oxypeucedanin hydrate (3), bergapten (5-methoxypsoralen) and six other known compounds, were isolated. Their structures were characterized basing on spectroscopic methods and chemical evidence. Compounds (1-3) were analyzed for their antimicrobial activity. Ficusamide (1) showed wick activity (minimal inhibitory concentration (MIC)=312.5 µg/mL) against Escherichia coli, while the furanocoumarins (2) and (3) showed significant activity (MIC=9.76 µg/mL) against Bacillus cereus, Candida albicans and Microsporum audouinii.

Chemical constituents from fruits of Cnestis ferruginea Vahl ex. DC (Connaraceae) and evaluation of their anticholinesterase and antiradical activities.

The phytochemical investigation of the DCM/MeOH (1:1) extract of the fruits of Cnestis ferruginea led to the isolation and characterization of one new quinic acid derivative, ferruginoic acid (1), together with six known compounds 2-7. Compounds 3-7 were reported for the first time from this species. The structures of compounds 1-7 were elucidated on the basis of 1 D and 2 D NMR spectroscopic data, mass spectrometry and by comparison of spectroscopic data with those from the literature. The anticholinesterase (AChE and BChE) activity and DPPH free radical scavenging assay of compounds 1, 3, 4 and 7 were evaluated. Ferruginoic acid (1) exhibited moderate anticholinesterase activity with IC50 value of 36.18 ± 1.78 µg/mL against AChE. Compounds 3, 4 and 7 showed high activity against free radical (DPPH•) scavenging assay (DPPH) with IC50 values 40.09 ± 0.96 µg/mL, 61.70 ± 0.78 µg/mL and 41.87 ± 0.62 µg/mL respectively. These results indicate that C. ferruginea and its constituents could be employed in the management of Alzheimer's disease.

Cytotoxic and genotoxic properties of artathomsonine, a new oxoberberine alkaloid from Artabotrys thomsonii (annonaceae).

A phytochemical investigation of the liana of Artabotrys thomsonii led to the isolation of a new oxoberberine alkaloid, 2,10-dihydroxy-3,9-dimethoxy-8-oxo-protoberberine (7), along with six known compounds. Their chemical structures were elucidated by 1 D and 2 D NMR spectroscopic methods and HRESI-MSn data analysis. Compounds 4 and 7 were selected for further in vitro investigations. In accordance with expectations from their chemical structures, compounds 7 and 4 showed a clear antioxidant activity in a cell-free assay, with compound 7 being 7-fold more active than 4. Cytotoxicity, cytostatic and genotoxic effects only occurred at high micromolar concentrations of 50 µM or more. Compound 7 was slightly less effective than compound 4. A low micromolar concentration of 10 µM did not cause any damaging cellular effects but showed potential for a protection against the micronucleus-inducing effect of reactive oxygen species hydrogen peroxide, although not to a significant extent.

A new cytotoxic indole alkaloid from Tabernaemontana inconspicua stapf.

One new indole alkaloid derivative, 5,6-dioxo-11-hydroxy voacangine (1) together with four known compounds (2-5), were isolated from the fruits of Tabernaemontana inconspicua Stapf (Apocynaceae). Their structures were determined using 1D and 2D NMR, HRESI-MS, and a comparison with the literature. The new compound was found to be cytotoxic on human breast cancer MDA-MB 231 cells with IC50 values of 3.35 µM and 2.19 µM after 24 and 48 hours, respectively.

Erymildbraedin A and B, two novel cytotoxic dimethylpyrano-isoflavones from the stem bark of Erythrina mildbraedii: evaluation of their activity toward endocrine cancer cells.

Two new dimethylpyrano-isoflavones, named erymildbraedin A (4) and B (5), were isolated from the stem bark of the Cameroonian medicinal plant Erythrina mildbraedii, along with four known ones, the linear congeners, scandenone (1), erysenegalinsein M (2), 5,4'-dihydroxy-2'-methoxy-8-(3,3-dimethylallyl)-2'',2''-dimethylpyrano[5,6:6,7]isoflavone (3), and the angular isoflavone eryvarin B (6), and two other compounds, fraxidin and scoparone. Their structures were elucidated by the usual spectroscopic methods and isoflavone effects on the growth of human breast, prostate, and endometrial adenocarcinoma cells were determined. Isoflavones 1, 3, and 6 strongly inhibited the growth of all three cell lines, supporting the notion that a non-oxidized isoprenyl group at C-8 is requisite for cytotoxic activity.

Antimicrobial and antioxidant effects of phenolic constituents from Klainedoxa gabonensis.

Bioassay-guided fractionation of the methanol extract of the stem bark of Klainedoxa gabonensis Pierre ex Engl. (Irvingiaceae) afforded 12 compounds, namely, ellagic acid (1), ellagic acid 3,3'-dimethylether (2), gallic acid (3), methyl gallate (4), lupeol (5), ß-amyrin (7), erythrodiol (8), oleanolic acid (9), betulinic acid (6), hederagenin (10), bayogenin acid (11), and stigmasterol-3-O-ß-d-glucopyranoside (12). Compounds 1-3 and 7-12 were isolated for the first time from this genus. The structures were established on the basis of 1D/2D NMR experiments and mass spectrometric data. Crude extract, fractions (A, B, C and D) and pure compounds were tested for their antimicrobial activity using paper disk agar diffusion assay. The test delivered a range of low to high activities for phenolic compounds 1-4, low or missing activities for terpenoid compounds 5-11, and impressive very high antibacterial/antifungal values for two fractions C and D probably due to synergistic effects of compounds. The broth microdilution assay revealed MICs of 15.4-115.1 µg/mL for phenolic compounds, MICs higher than 1 mg/mL for terpenoids and MICs of 4.5-30.3 µg/mL for fractions C and D. The determination of the radical scavenging activity using 1,1-diphenyl-2-picrylhydrazyl (DPPH) assay gave high antioxidant values for the methanol extract and fraction D (IC(50) 10.45 and 5.50 µg/mL) as well as for the phenolic compounds 1-4 (IC(50) 45.50-48.25 mM) compared to the standard 3-t-butyl-4-hydroxyanisole (BHA) (IC(50) 44.20 mM).

Triterpenoids with antimicrobial activity from Drypetes inaequalis.

The air-dried stems and ripe fruit of Drypetes inaequalis Hutch. (Euphorbiaceae) were studied. Four triterpene derivatives, characterized as lup-20(29)-en-3beta,6alpha-diol, 3beta-acetoxylup-20(29)-en-6alpha-ol, 3beta-caffeoyloxylup-20(29)-en-6alpha-ol and 28-betad-glucopyranosyl-30-methyl 3beta-hydroxyolean-12-en-28,30-dioate along with 10 known compounds were isolated from the whole stems. One triterpene, characterized as 3alpha-hydroxyfriedelan-25-al along with six known compounds were isolated from the ripe fruit. Their structures were established on the basis of spectroscopic analysis and chemical evidence. The triterpenes were tested for antimicrobial activity against some Gram-positive and Gram-negative bacteria, and two of them appeared to be modestly active.

New friedelane triterpenoids with antimicrobial activity from the stems of Drypetes paxii.

Two new friedelane-type triterpenes named 12alpha-hydroxyfriedelane-3,15-dione and 3beta-hydroxyfriedelan-25-al, together with six known compounds were isolated from the stems of Drypetes paxii Hutch. (Euphorbiaceae). Their structures were established on the basis of conventional 1 dimensional (1D) NMR methods, 2D shift-correlated NMR experiments and mass spectra. The five friedelane-type triterpene derivatives and one olean-12-ene triterpene saponin were tested for antimicrobial activity against some gram-positive and gram-negative bacteria, and they appeared to be modestly active.

Oligoamide, a new lactam from the leaves of Angylocalyx oligophyllus.

A new lactam, oligoamide (1), along with three known compounds (2-4), stigmasterol-3-O-ß-D-glucopyranoside (2), formononetin (3) and (-)-pinitol (4) were isolated from the CH2Cl2/CH3OH (1:1) extract of the leaves of Angylocalyx oligophyllus by chromatographic separation. Their structures were elucidated on the basis of spectroscopic analysis (UV, IR, MS, 1D, and 2D NMR). Compound 1 was found to have weak antioxidant and urease inhibitory potential.

Oxidative burst inhibitory and cytotoxic indoloquinazoline and furoquinoline alkaloids from Oricia suaveolens.

Two new ß-indoloquinazoline alkaloids, orisuaveoline A (1) and orisuaveoline B (2), two new furoquinoline alkaloids, quinosuaveoline A (5) and quinosuaveoline B (6), and 12 known compounds were isolated from Oricia suaveolens. The structures of the new compounds were deduced by spectroscopic studies. The absolute configuration of nkolbisine (4) was also determined. Compounds 2, 3, 6-8, 10, and 14 were evaluated for oxidative burst inhibitory activity in a chemoluminescence assay and for cytotoxicity against A549 lung carcinoma cells.

alpha-Glucosidase inhibitory constituents from stem bark of Terminalia superba (Combretaceae).

The CH(2)Cl(2)/CH(3)OH (1/1) extract of the dried stem bark of Terminalia superba afforded two compounds, (7S,8R,7'R,8'S)-4'-hydroxy-4-methoxy-7,7'-epoxylignan and meso-(rel 7S,8R,7'R,8'S)-4,4'-dimethoxy-7,7'-epoxylignan along with 11 known compounds. The structures of the compounds were established by analysing the spectroscopic data and also comparing it with the data of previously known analogues. All the isolated compounds were evaluated for their glycosidase inhibition activities. Gallic acid and methyl gallate showed significant alpha-glucosidase inhibition activity.

Secondary metabolites from Aspergillus japonicus CAM231, an endophytic fungus associated with Garcinia preussii.

Chemical investigation of Aspergillus japonicus CAM231, isolated from the leaf of Garcina preussii collected in Cameroon, yielded two new compounds; one pyrone derivative, hydroxy neovasinin (1) and one phenol derivative, asperolan (2), together with two known compounds neovasifurarone B (3) and variecolin (4). The structures of the two new compounds were established using intensive NMR spectroscopy and HRMS spectra in comparison with data found in literature. The structure of compound 1 was confirmed by single-crystal X-ray crystallographic analysis in combination with NOESY experiment. The new compounds were screened for their cytotoxic and antibacterial properties; however, the tested compounds displayed no significant activities.

Alkaloids from Oriciopsis glaberrima Engl. (Rutaceae).

Two alkaloid derivatives, oriciacridone A and B, were isolated from the stem bark of Oriciopsis glaberrima (Rutaceae). The structures were elucidated by a detailed spectroscopic analysis. The extract exhibited in vitro significant antimicrobial activity against a range of micro-organisms.

Phytochemistry and pharmacology of the genus Drypetes: A review.

AIMS: Traditional medicinal use of species of the genus Drypetes is widespread in the tropical regions. The aim of this review is to systematically appraise the literature available to date on phytochemistry, ethnopharmacology, toxicology and bioactivity (in vitro and in vivo) of crude extracts and purified compounds. ETHNOPHARMACOLOGICAL RELEVANCE: Plants of the genus Drypetes (Putranjivaceae) are used in the Subsaharan African and Asian traditional medicines to treat a multitude of disorders, like dysentery, gonorrhoea, malaria, rheumatism, sinusitis, tumours, as well as for the treatment of wounds, headache, urethral problems, fever in young children, typhoid and several other ailments. Some Drypetes species are used to protect food against pests, as an aphrodisiac, a stimulant/depressant, a rodenticide and a fish poison, against insect bites, to induce conception and for general healing. This review deals with updated information on the ethnobotany, phytochemistry, and biological activities of ethnomedicinally important Drypetes species, in order to provide an input for the future research opportunities. METHODS: An extensive review of the literature available in various recognized databases e.g., Google Scholar, PubMed, Science Direct, SciFinder, Web of Science, www.theplantlist.org and www.gbif.org, as well as the Herbier National du Cameroun (Yaoundé) and Botanic Gardens of Limbe databases on the uses and bioactivity of various species of the Drypetes was undertaken. RESULTS: The literature provided information on ethnopharmacological uses of the Subsaharan African and Asian species of the genus Drypetes, e.g., Drypetes aubrévillii, D. capillipes, D. chevalieri, D. gerrardii, D. gossweileri, D. ivorensis, D. klainei, D. natalensis, D. pellegrini (all endemic to Africa) and D. roxburghii (Asian species), for the treatment of multiple disorders. From a total of 19 species, more than 140 compounds including diterpenes, sesquiterpenes, triterpenes (friedelane, oleanane, lupane and hopane-type), flavonoids, lignans, phenylpropanoids and steroids, as well as some thiocyanates, were isolated. Several crude extracts of these plants, and isolated compounds displayed significant analgesic, anthelmintic, antidiabetic, anti-emetic anti-inflammatory, antioxidant, antiparasitic, central nervous system depressant, cytotoxic, and insecticidal activities both in vitro and in vivo. Some toxicities associated with the stem, bark, seed and leaf extracts of D. roxburghii, and the flavonoid, amentoflavone, isolated from the stem extract of D. littoralis as well as D. gerrardii, were confirmed in the animal models and in the rat skeletal myoblast cells assays. As a consequence, traditional medicine from this genus should in future be applied with care. CONCLUSIONS: Plants of this genus have offered bioactive samples, both from crude extracts and pure compounds, partly validating their effectivity in traditional medicine. However, most of the available scientific literatures lacks information on relevant doses, duration of the treatment, storage conditions and positive controls for examining bioefficacy of extract and its active compounds. Additional toxicological studies on the species used in local pharmacopeia are urgently needed to guarantee safe application due to high toxicity of some crude extracts. Interestingly, this review also reports 10 pimarane dinorditerpenoids structures with the aromatic ring C, isolated from the species collected in Asia Drypetes littoralis (Taiwan), D. perreticulata (China), and in Africa D. gerrardii (Kenya), D. gossweileri (Cameroon). These compounds might turn out to be good candidates for chemotaxonomic markers of the genus.

Anticancer and antibacterial secondary metabolites from the endophytic fungus Penicillium sp. CAM64 against multi-drug resistant Gram-negative bacteria.

BACKGROUND: The emergence of multiple-drug resistance bacteria has become a major threat and thus calls for an urgent need to search for new effective and safe anti-bacterial agents. OBJECTIVES: This study aims to evaluate the anticancer and antibacterial activities of secondary metabolites from Penicillium sp., an endophytic fungus associated with leaves of Garcinia nobilis. METHODS: The culture filtrate from the fermentation of Penicillium sp. was extracted and analyzed by liquid chromatography-mass spectrometry, and the major metabolites were isolated and identified by spectroscopic analyses and by comparison with published data. The antibacterial activity of the compounds was assessed by broth microdilution method while the anticancer activity was determined by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. RESULTS: The fractionation of the crude extract afforded penialidin A-C (1-3), citromycetin (4), p-hydroxyphenylglyoxalaldoxime (5) and brefelfin A (6). All of the compounds tested here showed antibacterial activity (MIC = 0.50 - 128 µg/mL) against Gramnegative multi-drug resistance bacteria, Vibrio cholerae (causative agent of dreadful disease cholera) and Shigella flexneri (causative agent of shigellosis), as well as the significant anticancer activity (LC50 = 0.88 - 9.21 µg/mL) against HeLa cells. CONCLUSION: The results obtained indicate that compounds 1-6 showed good antibacterial and anticancer activities with no toxicity to human red blood cells and normal Vero cells.