Unusual Sesquiterpenes from Streptomyces olindensis DAUFPE 5622.

In nature, the vast majority of sesquiterpenes are produced by type I mechanisms, and glycosylated sesquiterpenes are rare in actinobacteria. Streptomyces olindensis DAUFPE 5622 produces the sesquiterpenes olindenones A-G, a new class of rearranged drimane sesquiterpenes. Olindenones B-D are oxygenated derivatives of olindenone A, while olindenones E-G are analogs glycosylated with dideoxysugars. 13C-isotope labeling studies demonstrated olindenone A biosynthesis occurs via the methylerythritol phosphate (MEP) pathway and suggested the rearrangement is only partially concerted. Based on the structures, one potential mechanism of olindenone A formation proceeds by cyclization of the linear terpenoid precursor, likely occurring via a terpene cyclase-mediated type II mechanism whereby the terminal alkene of the precursor is protonated, triggering carbocation-driven cyclization followed by rearrangement. Diphosphate hydrolysis may occur either before or after cyclization. Although a biosynthetic route is proposed, the terpene cyclase gene responsible for producing olindenones currently remains unidentified.

Extraction and isolation of cannabinoids from marijuana seizures and characterization by 1H NMR allied to chemometric tools.

Marijuana, a drug derived from the Cannabis sativa L. plant, is the world's most consumed illicit drug. In this paper, a total of 156 marijuana samples seized in the state of Espírito Santo (ES), Brazil were studied and analysed by proton nuclear magnetic resonance (1H NMR) spectroscopy to identify the major cannabinoids present. A crude extract of all samples was purified using high performance liquid chromatography so that these compounds could serve as reference substances. Nine fractions were obtained and analysed by 1H NMR and gas chromatography-mass spectrometry (GC-MS), with five presented cannabinoids. ∆9-THC (Δ9-trans-tetrahydrocannabinol), ∆9-THCA (∆9-tetrahydrocannabinolic acid), ∆8-THC (∆8-tetrahydrocannabinol), 11-hydroxycannabinol, CBV (cannabivarin), and CBN (cannabinol) were found, and their chemical structures were confirmed by GC-MS. The latter compound was obtained with high purity (≈100%), while the others were obtained as less complex mixtures with purity higher than 75% (except for Δ8-THC). Principal component analysis (PCA) was used on the 1H NMR spectra of the 156 samples, and it was found that the samples were grouped according to the months, differentiating into two groups (from July 2014 to January 2015 and from February 2015 to July 2015), where non-grouping was observed from four macro-regions of the ES state (North, Central, Metropolitan, and South). The chemical profile of the seized samples was correlated to the 1H NMR spectrum of an isolated CBN sub-fraction, in which the group formed by samples seized in the year 2015 presented lower CBN content in the chemical composition. From the PCA score plot, two groups of samples were confirmed using the partial least squares discriminant analysis and orthogonal projections to latent structures classification methods.

Endophytic Actinobacteria from the Brazilian Medicinal Plant Lychnophora ericoides Mart. and the Biological Potential of Their Secondary Metabolites.

Endophytic actinobacteria from the Brazilian medicinal plant Lychnophora ericoides were isolated for the first time, and the biological potential of their secondary metabolites was evaluated. A phylogenic analysis of isolated actinobacteria was accomplished with 16S rRNA gene sequencing, and the predominance of the genus Streptomyces was observed. All strains were cultured on solid rice medium, and ethanol extracts were evaluated with antimicrobial and cytotoxic assays against cancer cell lines. As a result, 92% of the extracts showed a high or moderate activity against at least one pathogenic microbial strain or cancer cell line. Based on the biological and chemical analyses of crude extracts, three endophytic strains were selected for further investigation of their chemical profiles. Sixteen compounds were isolated, and 3-hydroxy-4-methoxybenzamide (9) and 2,3-dihydro-2,2-dimethyl-4(1H)-quinazolinone (15) are reported as natural products for the first time in this study. The biological activity of the pure compounds was also assessed. Compound 15 displayed potent cytotoxic activity against all four tested cancer cell lines. Nocardamine (2) was only moderately active against two cancer cell lines but showed strong activity against Trypanosoma cruzi. Our results show that endophytic actinobacteria from L. ericoides are a promising source of bioactive compounds.

Mycoleptones A-C and polyketides from the endophyte Mycoleptodiscus indicus.

Three new azaphilones with an unusual methylene bridge, named mycoleptones A, B, and C (2, 4, and 5), were isolated from cultures of Mycoleptodiscus indicus, a fungus associated with the South American medicinal plant Borreria verticillata. Additionally, four known polyketides, austdiol (1), eugenitin (3), 6-methoxyeugenin (6), and 9-hydroxyeugenin (7), were also isolated. The structural characterization of compounds was carried out by nuclear magnetic resonance spectroscopy, high-resolution mass spectrometry, electronic circular dichroism spectroscopy, time-dependent density functional theory calculations, and X-ray crystallography. Compounds 1-9 were weakly active when tested in antileishmanial and cytotoxicity assays.

Chiral HPLC analysis of donepezil, 5-O-desmethyl donepezil and 6-O-desmethyl donepezil in culture medium: application to fungal biotransformation studies.

An high performance liquid chromatography (HPLC) method for the enantioselective determination of donepezil (DPZ), 5-O-desmethyl donepezil (5-ODD), and 6-O-desmethyl donepezil (6-ODD) in Czapek culture medium to be applied to biotransformation studies with fungi is described for the first time. The HPLC analysis was carried out using a Chiralpak AD-H column with hexane/ethanol/methanol (75:20:5, v/v/v) plus 0.3 % triethylamine as mobile phase and UV detection at 270 nm. Sample preparation was carried out by liquid-liquid extraction using ethyl acetate as extractor solvent. The method was linear over the concentration range of 100-10,000 ng mL(-1) for each enantiomer of DPZ (r ≥ 0.9985) and of 100-5,000 ng mL(-1) for each enantiomer of 5-ODD (r ≥ 0.9977) and 6-ODD (r ≥ 0.9951). Within-day and between-day precision and accuracy evaluated by relative standard deviations and relative errors, respectively, were lower than 15 % for all analytes. The validated method was used to assess DPZ biotransformation by the fungi Beauveria bassiana American Type Culture Collection (ATCC) 7159 and Cunninghamella elegans ATCC 10028B. Using the fungus B. bassiana ATCC 7159, a predominant formation of (R)-5-ODD was observed while for the fungus C. elegans ATCC 10028B, DPZ was biotransformed to (R)-6-ODD with an enantiomeric excess of 100 %.

Novel [6]-gingerol Triazole Derivatives and their Antiproliferative Potential against Tumor Cells.

BACKGROUND: Effective cancer treatment is a major public health challenge. The limitations of current therapies and their adverse effects reduce the efficacy of treatment, leading to significant mortality rates worldwide. Moreover, natural product chemistry occupies a prominent role in the search for new treatment alternatives, by contributing a spectrum of chemical structures that may potentially yield new bioactive compounds. The compound [6]-gingerol (1) is the main active substance in ginger (Zingiber officinale) and several studies have shown it to produce beneficial effects, including antitumor activity. OBJECTIVE: This work aims to obtain new gingerol derivatives with cytotoxic activity. METHODS: [6]-gingerol was isolated and its derivatives were produced using click chemistry, obtaining eight new compounds. All chemical structures were determined by means of IR, NMR and HRMS data, and cytotoxicity was evaluated in the HCT 116 (colon carcinoma) and MCF-7 (breast carcinoma) cell lines at concentrations of 5 µmol L-1 and 50 µmol L-1. RESULTS: At 50 µmol L-1, more than 70% inhibition of cell growth was achieved with compounds 2e, 2g against HCT 116, and 2b, 2d, 2e, 2f and 2g against MCF-7. CONCLUSION: The obtained compounds showed only moderate cytotoxic activity. However, the products with substituents occupying the meta position in relation to the triazole ring showed increased cytotoxic properties. The brominated compound (2g) showed the strongest activity, inhibiting cell proliferation by 87%.

Structural and biosynthetic studies on eremophilenols related to the phytoalexin capsidiol, produced by Botrytis cinerea.

A thorough study of the fermentation broth of three strains of Botrytis cinerea which were grown on a modified Czapek-Dox medium supplemented with 5 ppm copper sulphate, yielded five undescribed metabolites. These metabolites possessed a sesquiterpenoid (+)-4-epi-eremophil-9-ene carbon skeleton which was enantiomeric to that of the phytoalexin, capsidiol. The isolation of these metabolites when the fungus was stressed, suggests that they may be potential effectors used by B. cinerea to circumvent plant chemical defences against phytopathogenic fungi. The biosynthesis of these compounds has been studied using 2H and 13C labelled acetate.

Anti-Acanthamoeba castellanii activity of alkaloid-enriched extracts and lycorine from the Amaryllidaceae species

Abstract Free-living amoebae of the genus Acanthamoeba are the causative agents of granulomatous encephalitis and keratitis, severe human infections. Bioactive compounds from plants are recognized as an alternative source for the development of new drugs. The Amaryllidaceae is a botanical family able to synthesize a very specific and consistent group of biologically active isoquinoline-like alkaloids. The alkaloidal fractions from the Brazilian species Hippeastrum canastrense, H. diniz-cruziae, H. puniceum, and Crinum x amabile, along with the alkaloid lycorine, were investigated against Acanthamoeba castellanii. The in vitro assays were performed with distinct concentrations of lycorine and alkaloidal fractions, while the cell viability was evaluated by the MTT method upon MDCK cells. Chlorhexidine 0.02% was used as the positive control. The effect of alkaloid fractions was concentration dependent, and 2000 µg mL-1 of H. canastrense and H. diniz-cruziae provided a 100% inhibition. At concentrations of 250, 500, and 1000 µg mL-1, the H. diniz-cruziae alkaloidal fraction showed the lowest cytotoxic effect (5%-7%) and remarkable anti-amoebic activity, demonstrating values of IC50 285.61 µg mL-1, low cytotoxicity (5%-7%), and selectivity index (7.0). Taken together, the results are indicative of the great potential that the alkaloids from H. diniz-cruziae have as new candidates for anti-amoebicidal compounds

Genome Sequence of Streptomyces sp. Strain RTd22, an Endophyte of the Mexican Sunflower.

We report here the complete genome sequence of Streptomyces sp. strain RTd22, an endophytic actinobacterium that was isolated from the roots of the Mexican sunflower Tithonia diversifolia The bacterium's 11.1-Mb linear chromosome is predicted to encode a large number of unknown natural products.

Analysis of 16S rRNA and mxaF genes revealing insights into Methylobacterium niche-specific plant association.

The genus Methylobacterium comprises pink-pigmented facultative methylotrophic (PPFM) bacteria, known to be an important plant-associated bacterial group. Species of this group, described as plant-nodulating, have the dual capacity of producing cytokinin and enzymes, such as pectinase and cellulase, involved in systemic resistance induction and nitrogen fixation under specific plant environmental conditions. The aim hereby was to evaluate the phylogenetic distribution of Methylobacterium spp. isolates from different host plants. Thus, a comparative analysis between sequences from structural (16S rRNA) and functional mxaF (which codifies for a subunit of the enzyme methanol dehydrogenase) ubiquitous genes, was undertaken. Notably, some Methylobacterium spp. isolates are generalists through colonizing more than one host plant, whereas others are exclusively found in certain specific plant-species. Congruency between phylogeny and specific host inhabitance was higher in the mxaF gene than in the 16S rRNA, a possible indication of function-based selection in this niche. Therefore, in a first stage, plant colonization by Methylobacterium spp. could represent generalist behavior, possibly related to microbial competition and adaptation to a plant environment. Otherwise, niche-specific colonization is apparently impelled by the host plant.

Antibacterial, antifungal and cytotoxic activities exhibited by endophytic fungi from the Brazilian marine red alga Bostrychia tenella (Ceramiales)

Abstract Marine environment is one of the most important sources regarding natural products research. Besides, marine microorganisms have been denominated as a talented natural source for discovery of new leads. Although the association of macroalgae and fungi has been described regarding ecological issues, there is a lack of studies about marine seaweed endophytic fungi. In this context, the goal of this study was to evaluate cytotoxic, antifungal and antibacterial activities of endophytic fungi isolated from the Brazilian marine seaweed Bostrychia tenella (J.V. Lamouroux) J. Agardh (Ceramiales, Rhodophyta). Forty-five endophytic microorganism strains were isolated from B. tenella. Crude extracts and organic fractions of ten selected strains were obtained after growth in rice medium. Samples were evaluated for cytotoxicity, antifungal and antibacterial assays. Penicillium strains showed positive results in a diversity of assays, and other five strains were active in at least one test. In addition, cytochalasin D was isolated from Xylaria sp. This alga is composed of a microbiological potential, since its endophytic strains exhibited remarkable biological properties. Moreover, cytochalasin D isolation has confirmed chemical potential of marine endophytic strains. This is the first study in which cultured fungi isolates from the Brazilian macroalga B. tenella were evaluated concerning biological properties. Results corroborated that this species could be a pharmaceutical source from marine environment. Furthermore, Acremonium implicatum is being firstly described as marine endophyte and Xylaria sp., Trichoderma atroviride and Nigrospora oryzae as marine seaweed endophytes. Thus, this work reports the first study relating detailed isolation, cultivation and biological evaluation (cytotoxic, antifungal and antibacterial) of endophytes Penicillium decaturense and P. waksmanii from the Brazilian marine red alga B. tenella. We are also reporting the isolation of cytochalasin D, a known antitumor and antibiotic compound, from Xylaria sp. strain. Despite widespread prevalence in terrestrial and marine habitats, this present work describes the first occurrence of cytochalasin D as a metabolite from marine seaweed endophyte.

Analysis of 16S rRNA and mxaF genes reveling insights into Methylobacterium niche-specific plant association

The genus Methylobacterium comprises pink-pigmented facultative methylotrophic (PPFM) bacteria, known to be an important plant-associated bacterial group. Species of this group, described as plant-nodulating, have the dual capacity of producing cytokinin and enzymes, such as pectinase and cellulase, involved in systemic resistance induction and nitrogen fixation under specific plant environmental conditions. The aim hereby was to evaluate the phylogenetic distribution of Methylobacterium spp. isolates from different host plants. Thus, a comparative analysis between sequences from structural (16S rRNA) and functional mxaF (which codifies for a subunit of the enzyme methanol dehydrogenase) ubiquitous genes, was undertaken. Notably, some Methylobacterium spp. isolates are generalists through colonizing more than one host plant, whereas others are exclusively found in certain specific plant-species. Congruency between phylogeny and specific host inhabitance was higher in the mxaF gene than in the 16S rRNA, a possible indication of function-based selection in this niche. Therefore, in a first stage, plant colonization by Methylobacterium spp. could represent generalist behavior, possibly related to microbial competition and adaptation to a plant environment. Otherwise, niche-specific colonization is apparently impelled by the host plant.

Bioactive extracts and chemical constituents of two endophytic strains of Fusarium oxysporum

Ethyl acetate extracts of cultures grown in liquid Czapek and on solid rice media of the fungal endophyte Fusarium oxysporum SS46 isolated from the medicinal plant Smallanthus sonchifolius (Poepp.) H. Rob., Asteraceae, exhibited considerable cytotoxic activity when tested in vitro against human cancer cells. Chromatographic separation yielded anhydrofusarubin (1) and beauvericin (2) that were identified based on their ¹H and 13C NMR data. Compounds 1 and 2 showed the strongest cytotoxic activity against different cancer cell lines. Compound 2 also showed promising activity against Leishmania braziliensis. Hexanic extract of F. oxysporum SS50 grown on solid rice media also afforded a mixture of compounds that displayed cytotoxic activity against different cancer cell lines. Chemical analysis of the mixture of compounds, investigated by gas chromatography-mass spectrometry (GC-MS), showed that there was a predominance of methyl esters of fatty acids and alkanes.

Avaliação microbiológica e físico-química de méis de jataí (Tetragonisca angustula) e de Apis mellifera do Estado de Mato Grosso do Sul / Microbiological evaluation and physicist-chemistry of honeys of jatai (Tetragonisca angustula) and Apis mellifera of the State of Mato Grosso do Sul

As abelhas jataí produzem mel com propriedades diferentes das abelhas africanas, sendo muito apreciado pelo seu sabor peculiar e pelas propriedades a ele atribuídas, muito embora poucos dados se dispõem sobre as qualidades físico-químicas e biológicas desse mel. Assim, os objetivos do trabalho foram avaliar o grau de contaminação de amostras de méis de abelhas jataí e de Apis mellifera de diferentes regiões do Estado de Mato Grosso do Sul, determinar alguns parâmetros físico-químicos, os quais podem fornecer dados para diferenciar os dois tipos de méis, bem como apresentar alguma correlação com a possível e/ou sobrevivência de microrganismos. Os méis analisados apresentaram pouca contaminação microbiana e não se mostraram contaminados por coliformes fecais ou coliformes a 45ºC, indicando produção e manipulação em boas condições higiênico-sanitárias. Através dos valores médios da umidade, acidez e Brix, os méis de jataí mostraram-se mais aquoso e mais ácidos que os de Apis mellifera e essas variáveis foram consideradas importantes para a diferenciação desses dois tipos de méis.