Structural basis of cycloaddition in biosynthesis of iboga and aspidosperma alkaloids.

Cycloaddition reactions generate chemical complexity in a single step. Here we report the crystal structures of three homologous plant-derived cyclases involved in the biosynthesis of iboga and aspidosperma alkaloids. These enzymes act on the same substrate, named angryline, to generate three distinct scaffolds. Mutational analysis reveals how these highly similar enzymes control regio- and stereo-selectivity.

Rotenoids from Clitoria fairchildiana R. Howard (Fabaceae) seeds affect the cellular metabolism of larvae of Aedes aegypti L. (Culicidae).

Non-domesticated species may represent a treasure chest of defensive molecules which must be investigated and rescued. Clitoria fairchildiana R. Howard is a non-domesticated Fabacea, native from the Amazonian Forest whose seeds are exquisitely refractory to insect predation. Secondary metabolites from these seeds were fractionated by different organic solvents and the CH2Cl2 fraction (CFD - Clitoria fairchildiana dichloromethane fraction), as the most toxic to 3rd instar Aedes aegypti larvae (LC50 180 PPM), was subjected to silica gel chromatography, eluted with a gradient of CH2Cl2: MeOH and sub fractioned in nine fractions (CFD1 - CFD9). All obtained fractions were tested in their toxicity to the insect larvae. Two rotenoids, a 11α-O-ß-D-glucopyranosylrotenoid and a 6-deoxyclitoriacetal 11-O-n-glucopyranoside, were identified in the mixture of CFD 7.4 and CFD 7.5, and they were toxic (LC50 120 PPM) to 3rd instar Ae. aegypti larvae, leading to exoskeleton changes, cuticular detachment and perforations in larval thorax and abdomen. These C. fairchildiana rotenoids interfered with the acidification process of cell vesicles in larvae midgut and caused inhibition of 55% of V-ATPases activity of larvae treated with 80 PPM of the compounds, when compared to control larvae. The rotenoids also led to a significant increase in the production of reactive oxygen species (ROS) in treated larvae, especially in the hindgut region of larvae intestines, indicating a triggering of an oxidative stress process to these insects.

Antihypertensive Activity of the Alkaloid Aspidocarpine in Normotensive Wistar Rats.

The alkaloid Aspidocarpine was isolated from the bark of Aspidosperma desmanthum. Its structure was elucidated by the spectral data of 1H and 13C-NMR (1D and 2D) and high-resolution mass spectrometry (HRESIMS). The antihypertensive activity was investigated by intravenous infusion in Wistar rats. This alkaloid significantly reduced (p < 0.05) the systolic, median, and diastolic blood pressures of rodents, without causing motor incoordination and imbalance in the rotarod test. The results indicate that the alkaloid Aspidocarpine exerts its antihypertensive activity without causing sedation or the impairment of motor functions.

Chemical characterization and in vitro biological activity of Cymbopogon citratus extracts against Haemonchus spp. and Trichostrongylus spp. nematodes from sheep.

Medicinal plants have been the focus of several studies due to their nematicide properties which can be used to control nematodes in sheep. No study has examined the morphological effects of Cymbopogon citratus on nematodes. Thus, this study evaluated the chemical composition, nematicidal activity and effects of C. citratus extracts on the morphology of eggs and infective larvae (L3) of sheep. Aqueous and methanolic extracts and fractions of C. citratus were obtained and analysed in vitro. The C. citratus extracts were effective against Haemonchus spp. and Trichostrongylus spp. larvae and eggs. Ten fractions were obtained from C. citratus, six of which had high ovicidal activity at 1000 µg mL-1, and two fractions had high activity at all tested concentrations. The phytochemical analysis identified the presence of compounds such as terpenoids, various ketones, esters, and fatty acids. The ultrastructural analysis showed deformations of the cuticle and wilting along the body of the nematodes at all concentrations. The muscular layer, intestinal cells and the mitochondria profile showed damage compared to the typical pattern. Ultra-thin sections of eggs treated with methanolic fractions of C. citratus presented modifications. This study showed the biological activity and effects of C. citratus on the gastrointestinal nematodes in sheep.

Plants as Sources of Anti-Inflammatory Agents.

Plants represent the main source of molecules for the development of new drugs, which intensifies the interest of transnational industries in searching for substances obtained from plant sources, especially since the vast majority of species have not yet been studied chemically or biologically, particularly concerning anti-inflammatory action. Anti-inflammatory drugs can interfere in the pathophysiological process of inflammation, to minimize tissue damage and provide greater comfort to the patient. Therefore, it is important to note that due to the existence of a large number of species available for research, the successful development of new naturally occurring anti-inflammatory drugs depends mainly on a multidisciplinary effort to find new molecules. Although many review articles have been published in this regard, the majority presented the subject from a limited regional perspective. Thus, the current article presents highlights from the published literature on plants as sources of anti-inflammatory agents.

Alkene lactones from Persea fulva (Lauraceae): Evaluation of their effects on tumor cell growth in vitro and molecular docking studies.

The new alkene lactone, (3E)-5,6-dihydro-5-(hydroxymethyl)-3-docdecylidenefuran-3(4H)-one (1), named majoranolide B, and three alkene lactones known as majorenolide (2), majoranolide (3) and majorynolide (4) were obtained from the aerial parts of Persea fulva (Lauraceae). The structures were elucidated in light of extensive spectroscopic analysis, including 1D, 2D NMR (1H, 13C, 1H-1H-COSY, HMBC and HSQC) and HR-ESI-MS. These compounds were screened for their in vitro antiproliferative activity in rat C6 glioma and astrocyte cells using MTT assay and in silico by molecular docking against targets that play a central role in controlling glioma cell cycle progression. Majoranolide (3) is the most active compound with IC50 6.69 µM against C6 glioma cells, followed by the compounds 1 (IC50 9.06 µM), 2 (IC50 12.04 µM) and 4 (IC50 41.90 µM). The alkene lactones 1-3 exhibited lower toxicity in non-tumor cells when compared to glioma cells. Molecular docking results showed that majoranolide establishes hydrogen bonds with all targets through its α,ß-unsaturated-γ-lactone moiety, whereas the long-chain alkyl group binds by means of several hydrophobic bonds. In the present study, it can be concluded from the anti-proliferative activity of isolates against C6 glioma cells that lactone constituents from P. fulva could have a great potential for the control of C6 glioma cells.

In Vitro Anti-Toxoplasma gondii and Antimicrobial Activity of Amides Derived from Cinnamic Acid.

Most cinnamic acids, their esters, amides, aldehydes, and alcohols present several therapeutic actions through anti-inflammatory, antitumor, and inhibitory activity against a great variety of microorganisms. In this work, eight amines derived from cinnamic acid were synthesized and tested against host cells infected with Toxoplasma gondii and the bacteria Escherichia coli, Pseudomonas aeruginosa, Staphylococcus epidermidis, and three strains of Staphylococcus aureus. Compounds 3 and 4 showed the best result against intracellular T. gondii, presenting antiparasitic activity at low concentrations (0.38 and 0.77 mM). The antibacterial activity of these compounds was also evaluated by the agar microdilution method, and amides 2 and 5 had a minimum inhibitory concentration of 250 µg mL-1 against two strains of S. aureus (ATCC 25923 and bovine strain LSA 88). These also showed synergistic action along with a variety of antibiotics, demonstrating that amines derived from cinnamic acid have potential as pharmacological agents.

Two Novel Cycloartane-Type Triterpenes from Trichilia casaretti C. DC. (Meliaceae).

Two new triterpenes cycloartane-type, named 24-methylencycloartan-12-oxo-3ß,22α-diol and trichiliol, were isolated from the leaves of Trichilia casaretti C. DC. together with three known triterpenes­24-methylencycloart-3ß,22-diol, 22,25-dihydrocycloart-23(E)-en-3ß-ol, and 22(R)-hydroxycycloart-24-en-3-ol. These compounds were characterized on the basis of their spectral data, mainly 1D (¹H and 13C) and 2D NMR (¹H-¹H-COSY, ¹H-¹H-NOESY, HMQC, HSQC, and HMBC), and mass spectra (EI-MS and HR-ESI-MS), also involving comparison with data from the literature.

13C-NMR Spectral Data of Alkaloids Isolated from Psychotria Species (Rubiaceae).

The genus Psychotria (Rubiaceae) comprises more than 2000 species, mainly found in tropical and subtropical forests. Several studies have been conducted concerning their chemical compositions, showing that this genus is a potential source of alkaloids. At least 70 indole alkaloids have been identified from this genus so far. This review aimed to compile 13C-NMR data of alkaloids isolated from the genus Psychotria as well as describe the main spectral features of different skeletons.

Chemicals from Agave sisalana biomass: isolation and identification.

Agave sisalana (sisal) is known worldwide as a source of hard fibers, and Brazil is the largest producer of sisal. Nonetheless, the process of removing the fibers of the sisal leaf generates 95% waste. In this study, we applied chemical sequential steps (hydrothermal extraction, precipitation, liquid-liquid extraction, crystallization, SiO2 and Sephadex LH 20 column chromatography) to obtain pectin, mannitol, succinic acid, kaempferol and a mixture of saponins as raw chemicals from sisal biomass. The structural identification of these compounds was performed though spectrometric methods, such as Infrared (IR), Ultraviolet (UV), Mass spectrometry (MS) and Nuclear magnetic resonance (NMR). All the sisal chemicals found in this work are used by both the chemical and pharmaceutical industries as excipients or active principles in products.

Chemistry and biological activity of ramalina lichenized fungi.

Lichens are a form of symbiont between a fungus and an alga or cyanobacterium, which contains a wide variety of organic compounds with certain secondary metabolite classes typical of these organisms. The Ramalina genus has approximately 246 species distributed around the World, of which in this review approximately 118 species with published chemical or biological activity studies of extracts or isolated compounds were cited. From the 153 mentioned compounds, only 27 passed were tested for biological activity, being usnic acid the most studied compound and the one showing the best results in almost all in vitro tests performed, although other compounds also presented excellent results as antimicrobial, antitumor and anti-inflammatory agents, among others. Extracts of several species also presented significant results in performed biological tests, demonstrating the potential that these organisms have, in particular, the gender Ramalina, to produce bioactive molecules that can be used as a model for the production of pharmaceuticals.

A novel 3,9-(1,2,3-trioxocine)-type steroid of Rauia nodosa (Rutaceae).

A new natural product, a 3,9-(1,2,3-trioxocine)-type steroid, named rauianodoxy (6), was isolated from Rauia nodosa, together with five steroids: sistostenone (1), stigmastenone (2), sitosterol (3), stigmasterol (4) and ergosterol peroxide (5), one coumarin, O-geranylosthenol (7), and three alkaloids, N-methylflindersine (8), zantobungeanine (9) and veprissine (10). Compounds 5-8 were isolated for the first time in the genus Rauia. These compounds were characterized on the basis of their spectral data, mainly one and two-dimensional NMR, and mass spectra, also involving comparison with the literature data. Theoretical studies at the DFT level reveal structural parameters for the 1,2,3-trioxole bridge compatible with known structures containing a similar group.

Hirtinone, a Novel cycloartane-type triterpene and other compounds from Trichilia hirta L. (Meliaceae).

One novel triterpene cycloartane-type, named hirtinone (1), six protolimonoids - nilocitin (2), dihydronilocitin B (3), melianone epimers (4) and (5), piscidinol A (6) and melianone lactone (7), one tertranortriterpenoid, hirtin (8), and one sesquiterpene, spathulenol (9), were identified in the fruits of Trichilia hirta. The structures were established by 1D and 2D NMR (1H and 13C-NMR, DEPTQ, 1H-1H-COSY, 1H-1H-NOESY, HSQC and HMBC), high resolution mass spectroscopy (HR-ESI-MS) and infrared (IR) spectral data.

A new cyclodipeptide from Tetragonisca angustula honey active against Neospora caninum and in silico study.

A new cyclic natural compound formed by succinic acid and two alanine amino acid units was isolated from the Tetragonisca angustula honey extract. The chemical structure of 1 was established based on spectroscopic data analysis, including one- (1H and 13C NMR) and two-dimensional NMR techniques (1H-1H-COSY, HSQC and HMBC). A primary culture model previously infected with Neospora caninum was used to evaluate 1 for two time intervals (24 and 72 h), showing a reduction (40-56%) of the number of tachyzoites in the first 24 h and until 72 h, a dose-dependent reduction in parasite proliferation (25-50%). Glial cells treated with 1 did not demonstrate toxicity at concentrations up to 25 ug/mL. Treated and infected cultures showed an increase in NO when compared to control cells in 24 h and 72 h. In silico studies suggest that the new compound may affect DNA synthesis and impair -protein production.

¹H and ¹³C-NMR data of the simplest plumeran indole alkaloids isolated from Aspidosperma species.

Indole alkaloids are the chemotaxonomic markers of the Aspidosperma genera. Those that have the simplest plumeran skeleton are classified as the precursors of biosynthetic routes and the intermediates for several synthetic reactions. This work aims to review the ¹H and ¹³C-NMR data, up to 2011, describing the skeleton of 35 different plumeran indole alkaloids, from a group of 46 of them, and highlight the main spectral differences amongst them.

New hydroperoxycycloartane-complete chemical shifts assignment of 13C and 1H-and cytotoxicity evaluation of cycloartanes isolated from Trichilia casaretti.

New cycloartane, 22-hydroxy-25-hydroperoxycycloart-23E-en-3-one (1), along with six known analogues (2-7) and three steroids (8-10), were isolated from the leaves of Trichilia casaretti. Structures were elucidated mainly on the basis of the analysis of 1D and 2D NMR (1H and 13C) and HRESIMS spectroscopic data, involving comparison with data of the literature. The cytotoxic activities of 1-7 and 10 isolated compounds were also evaluated against human leukemia cell line Molt-4 (acute lymphoblastic) and exhibited good cytotoxic activity with IC50 values ranging from 10.62 to 21.14 µM.

Antiplasmodial and cytotoxic effects of the methanol extract, canthinone alkaloids, squalene- and protolimonoid-type triterpenes from Homalolepis suffruticosa roots.

ETHNOPHARMACOLOGICAL RELEVANCE: Different species of the Simaroubaceae family are used in traditional medicine to treat malaria. Among these is Homalolepis suffruticosa (syn. Simaba suffruticosa and Quassia suffruticosa), which is native to Central Brazil and popularly known as calunga. However, there is a lack of investigation concerning its antimalarial effects. AIM OF THE STUDY: To investigate the antiplasmodial and cytotoxic effects of the isolated metabolites and methanol extract from H. suffruticosa roots as well as to conduct the dereplication of this extract aiming to characterize its metabolic profile by UPLC-DAD-ESI-MS/MS. MATERIALS AND METHODS: Methanol extract of the H. suffruticosa roots and six isolated compounds were evaluated against chloroquine-resistant Plasmodium falciparum W2 strain by the PfLDH method and cytotoxicity in HepG2 cells by the MTT assay. Dereplication of the extract was performed by UPLC-DAD-ESI-MS/MS. RESULTS: The six isolated compounds disclosed high to moderate antiplasmodial activity (IC50 0.0548 ± 0.0083 µg/mL to 26.65 ± 2.40 µg/mL) and cytotoxicity was in the range of CC50 0.62 ± 0.33 µg/mL to 56.43 ± 2.54 µg/mL, while 5-metoxycantin-6-one proved to be the most potent constituent of the six assayed ones. The methanol extract of the roots showed high in vitro antiplasmodial activity (IC50 1.88 ± 0.56 µg/mL), moderate cytotoxicity (CC50 41.93 ± 2.30 µg/mL), and good selectivity index (SI = 22.30). Finally, C20 quassinoids and canthin-6-one alkaloids were putatively identified in the H. suffruticosa methanol extract by LC-MS. CONCLUSIONS: Taken together, the isolated compounds, mainly the 5-metoxycantin-6-one and the methanol extract from H. suffruticosa roots, disclose good antiplasmodial activity, supporting the ethnopharmacological history of the Simaroubaceae species as traditional antimalarial drugs.

Terpenoids from endophytic fungi.

This work reviews the production of terpenoids by endophytic fungi and their biological activities, in period of 2006 to 2010. Sixty five sesquiterpenes, 45 diterpenes, five meroterpenes and 12 other terpenes, amounting to 127 terpenoids were isolated from endophytic fungi.

Missing enzymes in the biosynthesis of the anticancer drug vinblastine in Madagascar periwinkle.

Vinblastine, a potent anticancer drug, is produced by Catharanthus roseus (Madagascar periwinkle) in small quantities, and heterologous reconstitution of vinblastine biosynthesis could provide an additional source of this drug. However, the chemistry underlying vinblastine synthesis makes identification of the biosynthetic genes challenging. Here we identify the two missing enzymes necessary for vinblastine biosynthesis in this plant: an oxidase and a reductase that isomerize stemmadenine acetate into dihydroprecondylocarpine acetate, which is then deacetoxylated and cyclized to either catharanthine or tabersonine via two hydrolases characterized herein. The pathways show how plants create chemical diversity and also enable development of heterologous platforms for generation of stemmadenine-derived bioactive compounds.