Trypanocidal activity of South American Vernonieae (Asteraceae) extracts and its sesquiterpene lactones.

Chagas disease is caused by Trypanosoma cruzi and affects the poorest population in the Americas. Fourteen plant extracts and seventeen sesquiterpene lactones from the tribe Vernonieae (Asteraceae), were evaluated for the first time against T. cruzi. Cytotoxicity employing HeLa cells was also assessed. The best results were obtained with leaves and flowers rinse extracts from Vernonanthura nebularum (E-1 and E-3) and Elephantopus mollis (E-11 and E-13), with IC50 values <2 µg/mL, being E-1 the most active (IC50 = 0.8 µg/mL). Additionally, these extracts displayed a good selectivity (SI > 10). The most active sesquiterpene lactones, isolated from the extracts, were 2 (2-methoxy-2,5-epoxy-8-methacryloxygermacra-3Z,11(13)-dien-6,12-olide) and 6 (2-ethoxy-2,5-epoxy-8-angeloxygermacra-3Z,11(13)-dien-6,12-olide) from V. nebularum and 12 (8α-methacryloxyhirsutinolide 13-O-acetate) from V. pinguis, with IC50 of 1.5, 2.1 and 2.0 µM, respectively. These compounds showed SI values >14, better than those of the reference drug nifurtimox. Plants living in South American ecosystems could become a potential source of trypanocidal agents.

Lipophilic 9,10-Dehydrofukinone Action on Pathogenic and Non-Pathogenic Bacterial Biofilms. Why Is This Main Volatile Metabolite in Senecio?

The effect of a natural sesquiterpene ketone, 9,10-dehydrofukinone (DHF), on pathogenic Staphylococcus aureus and Pseudomonas aeruginosa strains isolated from chronic infectious processes, was the focus of the present study. Lipophilic DHF produced important antibacterial synergistic effects in association with ciprofloxacin (CPX) against two biofilm-forming strains of S. aureus HT1 (FIC=0.21) and P. aeruginosa HT5 (FIC=0.05). Hence, this mixture constitutes an excellent strategy to combat these biofilm-producing bacteria that overexpress drug efflux pumps as a resistance mechanism. Additionally, a substantial rise in beneficial Lactobacillus biofilm biomass was determined as a very significant finding of this association. Particularly, a non-pathogenic biofilm increment of 119 % was quantified when the mixture was added to a probiotic L. acidophilus ATCC SD-5212 culture. A surface activity enhanced in 71 % with respect to untreated L. acidophilus culture was also generated by the DHF and CPX association, and therefore, a glycoprotein synthesis induction mediated by the mixture is discussed. The results obtained could help in the development of new selective antibiotics. From an ecological standpoint, the present study strongly suggests that DHF is a polyfunctional organic molecule produced with a high yield in Senecio punae that exerts a positive impact on a non-pathogenic plant bacterium L. plantarum CE105.

Insecticidal effects of Vernonanthura nebularum against two economically important pest insects.

Vernonanthura nebularum (Cabrera) H. Rob. (Asteraceae), an endemic species from the north of Argentina, is a rich source of elephantopus-type sesquiterpene lactones. These compounds have proved to be promising antiparasitic agents, but there is no report about their action against pest insects. In this work we studied for the first time the antifeedant and toxic effects of V. nebularum natural products against the fall army warm Spodoptera frugiperda Smith and the oviposition deterrent activity against the fruit fly Ceratitis capitata Wiedemann. As a result, we found that extracts, fractions composed of sesquiterpene lactones and pure sesquiterpene lactones altered larval feeding behavior in the food choice test. Nutritional parameters of S. frugiperda larvae were also affected. Fraction II (300 µg/g of diet.), containing compounds 1, 2 and 3, was the most toxic substance with 80% pupal mortality and wing malformations in adults. In oviposition deterrent experiments against Ceratitis capitata, we observed a moderate effect at 30 µg/cm2 of the test compound. The most active substances were the methanolic extract, dichloromethane subextract and lactone 2. According to our results, V. nebularum natural products could be used for maximizing the effectiveness and specificity in future insecticide design with specific or multiple target sites, while ensuring the economic and ecological sustainability, in addition to combat the increasing resistance rates developed by synthetic pesticides.

Constituents of the Argentine Liverwort Plagiochila diversifolia and Their Insecticidal Activities.

Three new compounds, a bicyclogermacrene (1) and two 2,3-secoaromadendrane esters (2 and 3), together with (13S)-13-hydroxylabda-8,14-diene (4), fusicogigantone B (5), 3α,14-diacetoxy-2-hydoxybicyclogermacrene (6), fusicogigantone A (7), neofuranoplagiochilal (8), plagiochiline B (9), furanoplagiochilal (10), trans-nerolidol, spathulenol, α-tocopherol, and (+)-globulol were isolated from an Argentine collection of Plagiochila diversifolia. Their structures were elucidated by extensive mono and bidimensional NMR studies. Compounds 4, 5, and 6, incorporated to the larval diet at 100 µg per g of diet, reduced the larval growth of Spodoptera frugiperda (Lepidoptera: Noctuidae) by 70 ± 25, 57 ± 23, and 33 ± 16%, respectively. Compounds 4 and 5 produced 70% and 60% larval mortality at early instars. The latter also showed antifeedant properties in the Choice Test, with a feeding ratio of 0.54 ± 0.16.

Squamocin, an annonaceous acetogenin, enhances naphthalene degradation mediated by Bacillus atrophaeus CN4 / Squamocin, una acetogenina annonácea, estimula la degradación de naftaleno mediada por Bacillus atrophaeus CN4

Squamocin belongs to a group of compounds called annonaceous acetogenins. They are secondary products of Annonaceae metabolism and can be isolated from Annona cherimolia seeds. This paper deals with the stimulation of biofilm formation of Bacillus atrophaeus CN4 by employing low squamocin concentrations to increase naphthalene degradation. Bacillus atrophaeus CN4, isolated from contaminated soil, has the ability to degrade naphthalene as the only source of carbon and energy. In the absence of additional carbon sources, the strain removed 69% of the initial concentration of naphthalene (approx. 0.2 mmol/l) in the first 12 h of incubation. The addition of squamocin in LB medium stimulated Bacillus atrophaeus CN4 biofilm formation and enhanced naphthalene removal. Squamocin (2.5 pg/ml) does not affect planktonic growth and therefore, the observed increases are solely due to the stimulation of biofilm formation.

Squamocin pertenece a un grupo de compuestos llamados acetogeninas annonáceas (ACG). Las ACG son productos secundarios del metabolismo de plantas de la familia Annonaceae y se pueden aislar a partir de semillas de Annona cherimola. Este artículo trata de la estimulación de la formación de biofilm de Bacillus atrophaeus CN4 mediante el empleo de bajas concentraciones de squamocin para optimizar la degradación de naftaleno. B. atrophaeus CN4, aislado de suelo contaminado, tiene la capacidad de emplear naftaleno como única fuente de carbono y energía. En ausencia de fuentes de carbono adicionales, la cepa degradó el 69% de la concentración inicial de naftaleno (aprox. 0,2 mmol/l) en las primeras 12h de incubación. La adición de squamocin en medio LB estimula la formación de biofilm y la remoción naftaleno de B. atrophaeus CN4. Squamocin (2,5 µg/ml) no afecta al crecimiento planctónico y, por lo tanto, los incrementos observados se deben únicamente a la estimulación de la formación de biofilm.

Squamocin, an annonaceous acetogenin, enhances naphthalene degradation mediated by Bacillus atrophaeus CN4.

Squamocin belongs to a group of compounds called annonaceous acetogenins. They are secondary products of Annonaceae metabolism and can be isolated from Annona cherimolia seeds. This paper deals with the stimulation of biofilm formation of Bacillus atrophaeus CN4 by employing low squamocin concentrations to increase naphthalene degradation. Bacillus atrophaeus CN4, isolated from contaminated soil, has the ability to degrade naphthalene as the only source of carbon and energy. In the absence of additional carbon sources, the strain removed 69% of the initial concentration of naphthalene (approx. 0.2mmol/l) in the first 12h of incubation. The addition of squamocin in LB medium stimulated Bacillus atrophaeus CN4 biofilm formation and enhanced naphthalene removal. Squamocin (2.5µg/ml) does not affect planktonic growth and therefore, the observed increases are solely due to the stimulation of biofilm formation.

A New Depigmenting-Antifungal Methylated Grindelane from Grindelia chiloensis.

The new methylated grindelane diterpenoid, 7ß-hydroxy-8(17)-dehydrogrindelic acid (1b), together with the known 7α-hydroxy-8(17)-dehydrogrindelic acid (2a), 6-oxogrindelic acid (3a), 4ß-hydroxy-6-oxo-19-norgrindelic (4a), 19-hydroxygrindelic acid (5a), 18-hydroxygrindelic acid (6a), 4α-carboxygrindelic acid (7a), 17-hydroxygrindelic acid (8a), 6α-hydroxygrindelic acid (9a), 8,17-bisnor-8-oxagrindelic acid (10a), 7α,8α-epoxygrindelic acid (11a), and strictanonic acid (12a) as methyl esters were obtained from an Argentine collection of Grindelia chiloensis (Cornel.) Cabrera. Their structures and relative configurations were established on the basis of spectroscopic analysis. CHCl3 extract from the aerial parts and their pure compounds were evaluated for their antifungal and depigmenting effects. Methyl ester derivative of 10a (10b) exhibited a remarkable mycelial growth inhibition against Botritis cinerea with an IC50 of 13.5 µg ml-1 . While the new grindelane 1b exerted a clear color reduction of the yellow-orange pigment developed by Fusarium oxysporum against UV-induced damage.

Volatiles from Subtropical Convolvulaceae That Interfere with Bacterial Cell-to-Cell Communication as Potential Antipathogenic Drugs.

Increasing chronic bacterial infections create an urgent need for new antimicrobial agents or strategies for their control. Targeting virulence is one of the alternative approaches to find new medicines to treat persistent infections due to bacteria with biofilm-phenotype which are more resistant to antibiotics than their planktonic counterparts having an extreme capacity for evading the host defences. A bioguided study of sixteen extracts from flowers and leaves of four subtropical Convolvulaceae species provided evidence of the occurrence of antipathogenic natural products active against Gram positive and negative bacteria. Particularly, volatile metabolites from Merremia dissecta creeper, a food and medicinal plant, were able to interfere with the Pseudomonas aeruginosa quorum sensing system by a strong decrease of N-acyl homoserine lactone (AHL) biosynthesis (63-75%), which attenuated the virulence factor expression like biofilm (55%) and elastase activity (up to 27%), key factors that enable the colonization and dissemination of the infection in the host. Control of the P. aeruginosa biofilm and the QS process by phytochemicals, such as (+) spathulenol, isolated from a bioactive extract of M. dissecta leaves would be a good strategy for the development of new and effective antipathogenic drugs.

Active sesquiterpene lactones against Leishmania amazonensis and Leishmania braziliensis.

Seventeen sesquiterpene lactones (SLs) isolated from five species of the tribe Vernonieae were evaluated for their in vitro activity against promastigotes of Leishmania amazonensis and Leishmania braziliensis. Additionally, a quantitative structure activity relationship has been made, since all these natural compounds were found to have potent to mild antileishmanial properties. The most active compounds against L. braziliensis were 16 and 17 (IC50 values 1.45 and 1.34 µM, respectively), followed by compound 15 with IC50 value of 1.60 µM against L. amazonensis. The three glaucolide-type SLs (4-6) were the least active against both parasites. The computational study allowed us to establish that lipophilicity and polarisability play an important role in the antiparasitic activity. This is the first report of the known germacradiendiolides 16 and 17 from Elephantopus mollis. The activity data of the compounds 1-17 assayed against Leishmania parasites are reported here for the first time.

Prenylated Acylphloroglucinols with Leishmanicidal Activity from the Fern Elaphoglossum lindbergii.

Purification of a diethyl ether extract of the Argentinian fern Elaphoglossum lindbergii afforded five new prenylated acylphloroglucinols, lindbergins E-I (1-5), of which two showed significant in vitro leishmanicidal activity against promastigotes of Leishmania braziliensis and L. amazonensis. The structures of compounds 1-5 were elucidated based on analysis of their spectroscopic data and comparison with values previously reported for other phloroglucinol derivatives isolated from plant species of the genera Hypericum, Dryopteris, and Elaphoglossum. Fragmentation and rearrangement patterns of prenylated acylphloroglucinols were analyzed, and some mechanisms were proposed to rationalize the peaks observed in the mass spectra of these natural products produced by EI and FAB. Compounds isolated from E. lindbergii show the opposite absolute configuration when compared to those reported from E. crassipes. Empirical evidence indicates that acylphloroglucinols carrying a prenylated acylfilicinic acid residue possess a high-amplitude configuration-dependent Cotton effect centered at 350-360 nm in their CD curves, from which the absolute configuration of the sole chiral center of the prenylated acylfilicinic acid moiety can be deduced.

A Puna Collection of Senecio punae, Main Source of a Versatile Eremophilane-type Ketone.

Senecio punae Cabrera (Asteraceae-Senecioneae) is an endemic species from Puna semi-desert region of Argentina.The major compound, 4p,5p-eremophil- 7(11)9-dien-8-one, also known as dehydrofukinone (D), was isolated from the diethyl ether extract (E) of the plant. The phytochemical constituents from S. punae are presented here for the first time, as well as bioassays employing D, a molecule with versatility to carry out many different biological activities. E and D showed acute molluscicidal activity against the bilharzia vector snail Biomphalaria peregrina with LD50 values of 68.6 and 16.7 pg/mL, respectively. D exerted moderate and strong effects against promastigotes of Leishmania amazonensis and L. braziliensis, with IC5s values of 34.3?1.2 and 9.93+0.17 Ag/mL, respectively. Antibacterial effects were also found. Diethyl ether extract (E) and dehydrofukinone (D) were slightly active against Pseudomonas aeruginosa and Staphylococcus aureus at 50 and 25 tg/mL, respectively. However, D and E improved the effectiveness of oxacillin at 6 ig/mL against S. aureus growth and its biofilm, but did not affect beneficial lactobacilli strains. Synergistic effects were also measured between D (23 gg/mL) and ciprofloxacin (0.25 pg/mL) against two P. aeruginosa strains (FIC index = 0.38), as well as against two S. aureus strains (FIC indexes = 0.38 and 0.25). Finally, when 250 pg of either dehydrofukinone (D) or diethyl ether extract (E) were incorporated into the diet of the pest Oryzaephillus surinamensis, an alteration in the feeding behavior of the insect (Repellency indexes = +60 and +10, respectively) was observed.

Natural sesquiterpene lactones enhance oxacillin and gentamicin effectiveness against pathogenic bacteria without antibacterial effects on beneficial lactobacilli.

This is a report on the synergistic interactions (SIs) between melampolide-type sesquiterpene lactones 1-8 from Acanthospermum hispidum DC., and oxacillin or gentamicin, against four pathogenic strains of Staphylococcus aureus and Enterococcus faecalis; two of them were multi-resistant strains obtained from chronic infectious processes. Our results showed that all associations of 1-8 with antibiotics (ATBs) are more effective than pure ATBs to control pathogenic strains of S. aureus and E. faecalis. The most relevant SIs were observed when the major lactone of A. hispidum, acanthospermal B [5], was combined with gentamicin (protein synthesis inhibitor) against an ex vivo culture of methicillin-resistant S. aureus SAR 1, displaying a significant MIC reduction in 5 (312.5 to 78.1 µg/mL), and gentamicin (120 µg/mL to 3 µg/mL). Compound 4 improved the antibiotic potency of oxacillin (cell wall synthesis inhibitor) against ampicillin-resistant E. faecalis (60 µg/mL to 1.5 µg/mL). It is important to remark that three beneficial lactobacilli were resistant to 1-8 and their mixtures with gentamicin or oxacillin in effective concentrations against pathogenic bacteria. Synergism between ATBs and phytochemicals is a therapeutically helpful concept to improve ATB efficacy and prevent resistance. The present results show that selective SIs occur between melampolides and gentamicin or oxacillin, and open a new field of research.

Penicillium commune metabolic profile as a promising source of antipathogenic natural products.

Penicillium is an important genus of ascomycetous fungi in the environment and in food and drug production. This paper aims to investigate statins and antipathogenic natural products from a Penicillium commune environmental isolate. Fractions (F1, F2, F3 and F4) were obtained from an ethyl acetate extract. Direct insertion probe/electron ionisation/ion trap detection mass spectrometry (MS and MS/MS) identified lovastatin (1) in F1, while GC-MS showed that 3-isobutylhexahydropyrrolo[1,2-a]pyrazine-1,4-dione (2) was the main constituent of F2 (49.34%). F4 presented 3 (16.38%) as an analogue of 2 and their known structures were similar to that of an autoinducer-signal. F1 produced a significant decrease in the Pseudomonas aeruginosa biofilms, which is the main cause of bacterial pathogenicity. F2 and F4 were effective against Staphylococcus aureus biofilms, but when F2 was associated with oxacillin, it showed an important activity against both bacteria. These novel results suggest that P. commune INTA1 is a new source of promising antipathogenic products.

Toxic effect of citrus peel constituents on Anastrepha fraterculus Wiedemann and Ceratitis capitata Wiedemann immature stages.

The toxicity of essential oils from the citrus peel has been proposed as the major resistance mechanism offered by citrus to fruit fly infestation. We evaluated the insecticidal activity of the ether extracts from the lemon (Citrus limon [L.] Burm.) and grapefruit (C. paradisi Macfadyen) peel as well as from limonene and citral against Anastrepha fraterculus (Wiedemann) and Ceratitis capitata (Wiedemann) immature stages. We also evaluated the toxicity of the extracts at two ripening stages. Extracts proved toxic to A. fraterculus egg and larvae. The lemon and grapefruit extracts showed the same toxicity in both fruit fly species. For A. fraterculus eggs, citral was more toxic than limonene; for larvae, they showed equal toxicity. Anastrepha fraterculus eggs were more sensitive than C. capitata eggs. In conclusion, we provide evidence of chemical resistance mechanisms that could account for the nonhost condition of lemon for A. fraterculus.

Insecticidal properties of annonaceous acetogenins and their analogues. Interaction with lipid membranes.

The interactions were studied by FTIR and DSC of the terminal lactone of annonaceous acetogenins (ACGs) and synthetic analogues, such as THF, with POPC bilayers, as well as the toxic effect produced by these compounds on Spodoptera frugiperda larvae. The aim of this work was to find a relationship between ACG insecticidal properties and the specific sites of interaction with lipid membranes. ACGs interact to different extents with the phosphate of lipid membranes and differences in the antisymmetric stretching of the phosphate groups were found in the presence of water that indicate water loss and further hydrogen bonding.The ACG tested produced more than 70% larval mortality. Rolliniastatin-1 (3) proved to have the most toxic effects (100%) on early larval instars when incorporated in the larval diet at a dose of 100 microg per g of diet. Additionally, it produced a significant decrease in growth rate (GR) and consumption index (CI), and reduced the efficiency with which larvae converted ingested food into biomass (ECI). The destabilization that occurs in the membrane due to dehydration around the phosphate groups caused by interaction with ACGs and their synthetic analogues would account for ACGs' insecticidal action.

Inhibition of quorum sensing in Pseudomonas aeruginosa by sesquiterpene lactones.

Six sesquiterpene lactones (SLs) of the goyazensolide and isogoyazensolide-type isolated from the Argentine herb Centratherum punctatum were evaluated on their ability to inhibit virulence factors of Pseudomonas aeruginosa ATCC 27853. Although compounds were not able to completely inhibit bacterial growth at 200µg/ml, the SLs do altered biofilm formation, elastase activity, and production of N-acyl-homoserinelactones (AHLs) which are known quorum sensing autoinducers at lower concentration. Compounds 2, 3, and 5 displayed significant inhibitory effects on P. aeruginosa biofilm formation at 0.5µg/ml being compound 3 (1.32µM) the most potent (42%). Compounds 2, 3, 4, 5 and 6, inhibited 39, 44, 42, 32 and 35% the production of AHLs at 100µg/ml and inhibited by more than 50% the elastase activity at 0.5µg/ml. Our results clearly indicated that sesquiterpene lactones are good candidates for the development of new antimicrobial agents acting not as bactericidal but as antipathogenic agents.

Distribution of drimane sesquiterpenoids and tocopherols in liverworts, ferns and higher plants: Polygonaceae, Canellaceae and Winteraceae species.

The liverwort, Porella vernicosa complex produces a very hot tasting polygodial, a drimane-type sesquiterpene dialdehyde. The same compound has been isolated from two ferns, Thelypteris hispidula and Blechnum fluviatile, as well as from the higher plants Polygonum hydropiper, P. hydropiper f. purpurascens (Polygonaceae), Cinnamosma, Caspicodendron, Canella and Warburgia species (Canellaceae), and Pseudowintera colorata, Tasmannia lanceolata, Drimys and Zygogynum species (Winteraceae). In addition, the liverworts and higher plants which elaborate polygodial and its related pungent drimane dials contain a small amount of alpha-tocopherol, gamma-tocopherol or delta-tocotrienol. The present paper gives the results of a comparative study on the drimane-type sesquiterpenoids in some liverworts, ferns and higher plants, and the role of tocopherols in these plant groups.

Acylphloroglucinols from Elaphoglossum crassipes: antidepressant-like activity of crassipin A.

Nine new terpenylated acylphloroglucinols, crassipins A-I, were isolated from the rhizomes and roots of the fern Elaphoglossum crassipes, and their structures were elucidated by analysis of spectroscopic data and chemical derivatization. The absolute configurations of some of the compounds were established by CD and VCD in combination with a quantum mechanical method. Crassipin A (1), the major acylphloroglucinol of the Et(2)O extract of E. crassipes, as well as its peracetylated derivative (8), displayed antidepressant-like activity in a mouse forced-swimming test when administered orally at a dose of 15 mg/kg.

Unusual terpenylated acylphloroglucinols from Dryopteris wallichiana.

Four unusual terpenylated acylphloroglucinols were isolated from the diethyl ether extract of the scales and rhizomes of the fern Dryopteris wallichiana together with the known compounds albaspidins AA and AB, and filixic acids ABA and ABB. Structures of the isolated compounds were established by extensive spectroscopic analysis and their absolute configuration at C-14″ was determined by comparing their CD spectra with those simulated for the respective isomers. Pure acylphloroglucinols displayed moderate in vitro nematocidal activity against L4 stage larvae of Nippostrongylus brasiliensis (LD50=22-121 µM).

Bioactivity of the essential oil of an argentine collection of Acanthospermum hispidum (Asteraceae).

The essential oil of an Argentine collection of the annual herb Acanthospermum hispidum DC. (Asteraceae), which contains the sesquiterpenoids beta-caryophyllene (35.2%), alpha-bisabolol (11.4%), and germacrene D (11.1%) as major constituents, displayed molluscicidal effects on an adult population of Biomphalaria peregrina Orbigny (LD50 37.8 microg/mL) and produced alteration of the feeding and oviposition behavior of the polyphagous insect Spodoptera frugiperda Smith when incorporated in the larval diet at 250 microg/g. In addition, a selective antibacterial activity was observed against pathogenic strains of Staphylococcus aureus and Enterococcus faecalis (MICs 62.5 and 125 microg/mL), but was inactive at 125 microg/mL against three beneficial lactic acid bacteria. Synergistic interactions were further validated by FIC index determination of the combination of the antibiotics oxacillin and gentamicin and the essential oil on the four pathogenic strains evaluated.