Semi-purified Fractions Research Articles

Evaluation of anti-inflammatory, antiangiogenic and antiproliferative activities of Arrabidaea chica crude extracts

Ethnopharmacological relevanceArrabidaea chica (Bignoniacea) has been used in popular medicine in Brazil to treat inflammation, skin diseases and leukemia. This work aimed to investigate the anti-inflammatory and antitumoral activities of the A. chica aqueous (AE) and ethanol (EE) extracts. Materials and methodsThe murine sponge model was used to evaluate the anti-inflammatory and antiangiogenic activities of AE and EE. Accumulation of neutrophil and macrophage in the implants were determined by assaying myeloperoxidase and N-acetyl-glucosaminidase activities and the neovascularization evaluated by the amount of hemoglobin present in the implant using the Drabkin method. The antitumoral activity was evaluated using the MTT colorimetric method against Jurkat, HL60 and MCF-7 cells. Semi-purified fractions F1-F4 from the EE extract were obtained by a liquid–liquid solvent extraction method and their in vitro anti-proliferative effects were also investigated. ResultsEthanol and aqueous extracts of A. chica decreased neutrophil accumulation and hemoglobin content in the sponge implants without altering the level of cytokines (IL-2, IL- 4, IL-5, IFN-γ, TNF-α and VEGF) and the albumin/globulin ratio in the serum of treated animals. There was no sign of toxicity (clinical, laboratory or histopathology). The ethanol extract presented antiproliferative activity (IC50 21.5–36.3µg/mL) against HL60 and Jurkat cell lineages and proapoptotic activity at 50µg/mL in HL60 cells. The fraction F1 also demonstrated significant antiproliferative activity (IC50 38.5µg/mL) and proapoptotic activity against HL60 cells in a dose dependent manner. ConclusionsAqueous and ethanol extracts of A. chica attenuate the inflammatory and angiogenic components of the subcutaneous fibrovascular tissue induced by the synthetic matrix in mice. In addition, the ethanol extract from Arrabidaea chica and its fraction F1 presented in vitro antiproliferative activity and could be useful for developing potential chemopreventive substances.

Piperamides fromPiper ottonoidesby NMR and GC-MS Based Mixture Analysis

The species Piper ottonoides Yuncker (Piperaceae), known as “joao-brandim”, is a shrub that occurs in the Brazilian Amazon rainforest. Its roots and leaves are used in traditional medicine as local anesthetic to treat toothache and sore throat. In this study, the structural characterization in mixture of isobutyl amides present in semi-purified fractions from fruits, leaves, stems and roots of P. ottonoides was achieved by employing gas chromatography-mass spectrometry (GC-MS) and a combination of nuclear magnetic resonance (NMR) techniques (1H, 1H-1H COSY, 1H-13C HSQC, 1H-13C HMBC, TOCSY and 1H-1H J-RES). The MS fragmentation patterns and the NMR chemical shifts, multiplicities, coupling constants and the signal correlations in the two-dimensional spectra were carefully analyzed also taking into account some key elements of differentiation among the compounds. The data set allowed identifying unequivocally the new amide N-isobutyl-7-(4’- methoxyphenyl)-2E,4E-heptadienamide, which we named ottonoidenamide as well as piperovatine and chingchengenamide A in all parts of the plant, the additional presence of pipercallosine and pipercallosidine in roots and of dihydropiperlonguminine in fruits and leaves. The presence of piperovatine and pipercallosine in P. ottonoides should be associated to its traditional use.

Enhanced haemagglutination and phytochemical evaluation suggest significant presence of glycoproteins (Lectins) in semi-purified fractions of eastern Nigeria Mistletoe, Loranthus micranthus Linn.

Enhanced haemagglutination and phytochemical evaluation suggest significant presence of glycoproteins (Lectins) in semi-purified fractions of eastern Nigeria Mistletoe, Loranthus micranthus Linn.

Metabolites derived from the tropical seagrass Thalassia testudinum are bioactive against pathogenic Labyrinthula sp

Temperate and tropical seagrasses are susceptible to wasting disease outbreaks caused by pathogenic protists of the genus Labyrinthula. Even though there is an increasing awareness of the environmental conditions that influence the etiology of seagrass-Labyrinthula disease dynamics, the biochemical basis of seagrass defense responses, in particular chemical defenses, is still vastly understudied. Using an in vitro bioassay, we provide evidence that previously characterized phenolic and potentially novel, undescribed non-phenolic metabolites derived from Thalassia testudinum Banks ex Konig exhibit anti-labyrinthulid activity. All phenolic compounds tested displayed dose-dependent behavior and selected combinations interacted synergistically. The flavone glycoside thalassiolin B was roughly 20–100 times more active than any phenolic acid tested. Based upon values reported in the literature, it was calculated that infected specimens of T.testudinum contain natural concentrations of phenolic acids that are consistently greater than what is required to inhibit Labyrinthula growth. This suggests that while there may be an ample supply of phenolic-based derivatives available to inhibit Labyrinthula growth, they may not be readily bio-accessible.Using a bioactivity-guided approach, a semi-purified chemical fraction from T. testudinum was found to contain anti-labyrinthulid activity. 1H NMR spectra for this fraction lacked aromatic hydrogen signals, suggesting that the bioactive compound was non-aromatic in nature. Furthermore, the LC–MS fragmentation patterns were suggestive of the presence of glycosylated natural products of an unknown structural class. This has the potential to provide a foundation for future chemical investigations.

Antioxidant and anti-inflammatory activities contribute to the prophylactic effect of semi-purified fractions obtained from the crude methanol extract of Muntingia calabura leaves against gastric ulceration in rats

Ethnopharmacological relevanceIn traditional medicine, the leaves, flowers, barks and roots of Muntingia calabura L. (Muntingiaceae) have been employed as a treatment for various ailments including dyspepsia and to relieve pain caused by gastritis and peptic ulcer disease. The methanolic extract of Muntingia calabura leaves (MEMC) has been proven in the previous study to possess significant antiulcer activity. In this study, we attempted to determine the prophylactic effect of the fractions obtained from MEMC against ethanol-induced gastric lesion in rats and the involvement of antioxidants and anti-inflammatory mediators. Materials and methodsThe MEMC was fractionated with petroleum ether (PEF), ethyl acetate (EAF) and distilled water (AQF). These fractions were investigated for possible antiulcer property using ethanol-induced gastric ulcer rat model. The rats were administered orally once daily with 8% Tween 80 (control), 100mg/kg ranitidine, or the fractions, in the doses of 100, 250, and 500mg/kg, for 7 days, followed by ulcer induction using absolute ethanol. The rats were euthanized; macroscopic and histological observations of the stomach were done. The ulcer area (UA) was determined and the percentage protection afforded by the fractions was calculated. The fractions were subjected to antioxidant studies including the superoxide and 2, 2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging, oxygen radical absorbance capacity (ORAC) and total phenolic content (TPC) assay. Involvement of nitric oxide (NO) and inflammatory mediators such as lipoxygenase (LOX) and xanthine oxidase (XO) were evaluated. Phytochemical screening and HPLC analysis of the fractions were also conducted. ResultsPre-treatment of PEF and EAF significantly (p<0.001) attenuated the gastric lesions as compared to the control group in a dose-dependent manner. On the other hand, 100 and 250mg/kg of AQF significantly (p<0.001) prevented the ulcer formation but at the highest dose (500mg/kg), AQF failed to significantly reduce the ulcer formation, showing a dose-independent antiulcerative effect of AQF. The histological evaluation supported the observed gastroprotective activity of PEF, EAF and AQF. All the fractions showed high superoxide and DPPH scavenging activity, meanwhile the EAF showed highest TPC followed by PEF and AQF. These fractions also significantly (p<0.05) inhibited the NO while maintaining the viability of the cells. EAF exhibited high inhibition towards both the LOX and XO enzymes, meanwhile PEF and AQF exerted high LOX inhibition but low XO inhibition. Phytochemical screening and HPLC profiling suggested the presence of flavonoid- and tannin based compounds in PEF and EAF. ConclusionIt can be concluded that the prophylactic effect of the fractions on gastric ulceration in rats is associated with its high antioxidant activity and its ability to effectively inhibit the inflammation mediators. Presence of several flavonoids and gallic acid explains the effectiveness of the fractions in affording protection against gastric damages.

Evaluation of anticandidal and antioxidant activities of phenolic compounds from Pyrostegia venusta (Ker Gawl.) Miers.

We have investigated the in vitro anticandidal and antioxidant activities of phenolic compounds from Pyrostegia venusta flower extracts. We used the HPLC technique to purify the flavonoid (quercetin-3-O-α-l-rhamnopyranosyl-(1→6)-β-d-galactopyranoside) and two phenylpropanoid glycosides (verbascoside and isoverbascoside); we evaluated the antimicrobial activity of the extracts against Candida strains (Candidaalbicans; Candidakrusei ATCC 6258; and the clinical isolate strains of Candida sp. C. albicans, C. krusei, Candidatropicalis, Candidaparapsilosis, and Candidaguilhermondii). The P. venusta flower extracts displayed antimicrobial and antioxidant activities. The semi-purified fraction of the P. venusta flower extract and the phenylpropanoid glycoside verbascoside exhibited activity similar to that of amphotericin B, which denoted that they are potentially applicable as natural antioxidant and anticandidal agents in the pharmaceutical industries.

Pharmacological evaluation of the semi-purified fractions from the soft coral Eunicella singularis and isolation of pure compounds.

BackgroundGorgonians of the genus Eunicella are known for possessing a wide range of pharmacological activities such as antiproliferative and antibacterial effect. The aim of this study was to evaluate the anti-inflammatory and gastroprotective effect of the organic extract and its semi-purified fractions from the white gorgonian Eunicella singularis and the isolation and identification of pure compound(s) from the more effective fraction.MethodsAnti-inflammatory activity was evaluated, using the carrageenan-induced rat paw edema test and in comparison to the reference drug Acetylsalicylate of Lysine. The gastroprotective activity was determined using HCl/EtOH induced gastric ulcers in rats. The purification of compound(s) from the more effective fraction was done by two chromatographic methods (HPLC and MPLC). The structure elucidation was determined by extensive spectroscopic analysis (1H and 13C NMR, COSY, HMBC, HMQC and NOESY) and by comparison with data reported in the literature.ResultsThe evaluation of the anti-inflammatory activity of different fractions from Eunicella singularis showed in a dependent dose manner an important anti-inflammatory activity of the ethanol fraction, the percentage of inhibition of edema, 3 h after carrageenan injection was 66.12%, more effective than the reference drug (56.32%). In addition, this ethanolic fraction showed an interesting gastroprotective effect compared to the reference drugs, ranitidine and omeprazol. The percentage of inhibition of gastric ulcer induced by HCl/ethanol in rats was 70.27%. The percentage of the reference drugs (ranitidine and omeprazol) were 65 and 87.53%, respectively. The purification and structure elucidation of compound(s) from this ethanolic fraction were leading to the isolation of five sterols: cholesterol (5α-cholest-5-en-3β-ol) (1); ergosterol (ergosta-5,22-dien-3β-ol) (2); stigmasterol (24-ethylcholesta-5,22-dien-3b-ol) (3); 5α,8α-epidioxyergosta 6,22-dien-3β-ol (4) and 3β-hydroxy-5α,8α-epidioxyergosta-6-ene (5); and one diterpenoid: palmonine D (6).ConclusionBased on data presented here, we concluded that diterpenoids and sterols detected in the ethanolic fraction can be responsible for its pharmacological activity.Electronic supplementary materialThe online version of this article (doi:10.1186/s40199-014-0064-7) contains supplementary material, which is available to authorized users.

Cistanches Herba reduces the weight gain in high fat diet-induced obese mice possibly through mitochondrial uncoupling

A semi-purified fraction isolated from Cistanches Herba (HCF1), was previously found to induce mitochondrial uncoupling in H9c2 cells and in rat hearts. We therefore hypothesized that HCF1 would produce weight loss effect against a high fat diet (HFD)-induced obesity. To test this hypothesis, a mouse model of HFD-induced obesity was established and the effects of HCF1 on normal diet (ND)-fed and HFD-fed mice were examined. In the study, long term HCF1 treatment produced weight reduction effect against HFD-induced obesity in male and female mice. The HCF1-induced weight loss was associated with improved insulin sensitivity in HFD-fed animals. To understand the action mechanism underlying the weight reduction effect afforded by HCF1, its effects on mitochondrial uncoupling was examined. A comparative study with cholestyramine (CT), a bile acid sequestrant, was also conducted. The findings demonstrated that HCF1-induced weight loss was likely mediated by the increase in energy consumption, probably via the induction of mitochondrial uncoupling in mouse skeletal muscle. Thus, our findings suggest the potential use of HCF1 to prevent obesity and the associated health consequences such as diabetes, cardiovascular diseases and metabolic syndrome.

Oncibauerins A and B, new flavanones from Oncidium baueri (Orchidaceae)

The phytochemical study of Oncidium baueri (Orchidaceae) afforded two new compounds (1 and 2), along with thirteen known structures (3–15). The chemical structures were established on the basis of spectral evidence and the new compounds, obtained as a mixture of two glycosylated flavanones were assigned by comprehensive spectroscopic analysis by 1D and 2D NMR spectroscopy and HRMS. Moreover, the cytotoxic activity of the crude extract (EBOB), semi-purified fractions (HEOB, CLOB, ACOB, BUOB and HMOB) and some isolated metabolites against ten human cancer cell lines (U251, UACC-62, MCF-7, NCI-ADR/RES, 786-0, NCI-H460, PC-3, OVCAR-3, HT29, K562) and the antiprotozoal activity against Trypanosoma cruzi and Leishmania amazonensis were evaluated. The CLOB fraction showed the best antitrypanosomal and antileishmanial activities, as well as considerable cytotoxicity against cancer cell lines UACC-62 (melanoma) and K562 (leukemia) with GI50 values of 6.83μg/mL and 0.5μg/mL, respectively. Furthermore, BUOB and HMOB fractions showed selectivity for UACC-62 (melanoma) with GI50<0.01μg/mL and the compounds 11, 13 and 14 had inhibited cell growth of almost cancer cell lines tested with GI50 values close to 30μg/mL.

Pseudomonas aeruginosa elastase disrupts the cortisol-binding activity of corticosteroid-binding globulin.

The serine protease inhibitor (SERPIN) family member corticosteroid-binding globulin (CBG) is the main carrier of glucocorticoids in plasma. Human CBG mediates the targeted release of cortisol at sites of inflammation through cleavage of its reactive center loop (RCL) by neutrophil elastase. The RCLs of SERPIN family members are targeted by diverse endogenous and exogenous proteases, including several bacterial proteases. We tested different bacteria for their ability to secrete proteases that disrupt CBG cortisol-binding activity, and characterized the responsible protease and site of CBG cleavage. Serum CBG integrity was assessed by Western blotting and cortisol-binding capacity assay. Effects of time, pH, temperature, and protease inhibitors were tested. Proteolytically active proteins from bacterial media were purified by fast protein liquid chromatography, and the active protease and CBG cleavage sites were identified by mass spectrometry. Among the bacteria tested, medium from Pseudomonas aeruginosa actively disrupted the cortisol-binding activity of CBG. This proteolytic activity was inhibited by zinc chelators and occurred most efficiently at pH 7 and elevated physiological temperature (ie, 41°C). Mass spectrometric analysis of a semi-purified fraction of P. aeruginosa media identified the virulence factor LasB as the responsible protease, and this was confirmed by assaying media from LasB-deficient P. aeruginosa. This metalloprotease cleaves the CBG RCL at a major site, distinct from that targeted by neutrophil elastase. Our results suggest that humoral responses to P. aeruginosa infection are influenced by this pathogen's ability to secrete a protease that promotes the release of the anti-inflammatory steroid, cortisol, from its plasma transport protein.

Pseudomonas aeruginosa produces secondary metabolites that have biological activity against plant pathogenic Xanthomonas species

The bioactivity of compounds produced by Pseudomonas aeruginosa (LN strain), against three Xanthomonas species was investigated under greenhouse conditions and using electron microscopy. Chromatographic fractions EAP, VLC3, VLC4, VLC3d and VLC4f were tested against Xanthomonas axonopodis pv. malvacearum, X. axonopodis pv. phaseoli and X. axonopodis pv. citri by disc diffusion. Fractions with antibiotic activity were tested in vivo under greenhouse conditions and their bioactivity was evaluated. Scanning electron microscopy showed that VLC4f affects biofilm formation while VLC4f and VLC3d both affect cell morphology. The semi-purified fractions controlled bacterial diseases caused by Xanthomonas spp. when sprayed on plants under greenhouse conditions. The VLC4f fraction showed superior results in disease management, reducing the number of lesions on cotton and orange leaves by 94%, and reducing disease severity in bean leaves by 73%. The data suggest that the fractions were effective and have potential as an alternative to conventional bactericides in the control of plant diseases caused by Xanthomonas sp under greenhouse conditions.

SEPARATION AND PURIFICATION OF ANTIOXIDANT GLYCOGLYCEROLIPIDS FROM FRUIT OF CUCURBITA MOSCHATA BY HIGH-SPEED COUNTER-CURRENT CHROMATOGRAPHY COUPLED WITH PREPARATIVE HIGH-PERFORMANCE LIQUID CHROMATOGRAPHY

Glyceroglycolipids, widely distributed in edible plants, possess potent antithrombotic, antiviral, antitumor, and anti-inflammatory activities. In this paper, a simple method based upon high-speed counter-current chromatography (HSCCC) coupled with preparative high-performance liquid chromatography (Pre-HPLC) was developed for the separation and purification of antioxidant glycoglycerolipids from the fruits of Cucurbita moschata (pumpkin). HSCCC separation was performed on a two-phase solvent system composed of dichloromethane–light petroleum (60–90°C)–ethanol–H2O (6:2:4:4, v/v). Semi-purified peak fractions from HSCCC separation were further purified by Pre-HPLC to yield six glycoglycerolipid monomers with purities of over 98%. Their structures were elucidated by electrospray ionization mass spectrometry (ESI-MS), 1H-NMR, and 13C-NMR. Their antioxidant activities in vitro were also evaluated through 1,1-diphenyl-2-picrylhydrazyl assays. Tetrasaccharide-containing glycoglycerolipids exhibited stronger antioxidant activity compared to that of butylated hydroxytoluene, and it can be assumed to be potentially useful as a safe antioxidant in food processing industries. The results indicate that the method is simple, fast, and efficient.

Assessment of the spatial variability of phenolic contents and associated bioactivities in the invasive alga Sargassum muticum sampled along its European range from Norway to Portugal

Sargassum muticum, an invasive brown macroalga presently distributed along European Atlantic coasts from southern Portugal to the south coast of Norway, was studied on a large geographical scale for its production of phenolic compounds with potential industrial applications and their chemical and biological activities. S. muticum can produce high biomass in Europe, which could be exploited to supply such compounds. S. muticum was collected in Portugal, Spain, France, Ireland and Norway (three sites/country) to examine the effect of the latitudinal cline and related environmental factors. Assays focused particularly on polyphenols and their activities. Crude acetone–water extracts were purified using solid phase extraction (SPE) and antioxidant and antimicrobial activities of crude extracts and semi-purified fractions measured. Total phenolic content was assessed by colorimetric Folin–Ciocalteu assay and reactive oxygen species activities by 2,2-diphenyl-1-picrylhydrazyl, reducing power, β-carotene bleaching method and xanthine oxidase assay. Antibacterial activities were tested on terrestrial and marine strains to evaluate potential use in biomedical and aquaculture fields. Purified active phlorotannins, isolated by SPE, were identified using NMR. Phenolic contents differ clearly among countries and among sites within countries. Quality did not change between countries, however, although there were some slight differences in phlorethol type. Additionally, some fractions, especially from the extreme north and south, were very active. We discuss this in relation to environmental conditions and the interest of these compounds. S. muticum represents a potential natural source of bioactive compounds and its collection could offer an interesting opportunity for the future management of this species in Europe.

Antimicrobial activity of crude and semi-purified fractions of Warburgia ugandensis against some pathogens

Antimicrobial activity of crude and semi-purified fractions of Warburgia ugandensis against some pathogens

A TRIGLYCERIDE FROM LUPINUS ALBUS L. SEED WITH BIO-STIMULATORY PROPERTIES

In vitro and in vivo bio-stimulatory activity of a Lupinus albus ‘Bethuana White’ seed suspension (SS) was previously documented using vegetable crops. This was repeated for two grain crops, maize and wheat, in an attempt to provide a rationale for the isolation and identification of the active compound(s) involved. In all cases, besides negative controls, the response to treatment with SS was compared to that of a commercial bio-stimulant, ComCat®. At an optimum concentration of 5 mg/L SS significantly enhanced seedling growth and yield in both crops. Subsequently, activity directed liquid-liquid extraction of ground L. albus seed resulted in an ethyl acetate extract containing most of the bio-stimulatory activity. Two in vitro bio-assay procedures were employed to appraise the in vitro bio-stimulatory properties of semi-purified fractions as well as pure compounds contained in it. The highly active ethyl acetate extract was fractionated further by means of adsorption column chromatography resulting in eighty five fractions that were combined into twelve on the basis of similarities in thin layer chromatography profiles. Four of these fractions showed significant bio-stimulatory activity and were pooled. From this one compound was isolated, purified and identified as glyceryl trilinoleate by means of nuclear magnetic resonance spectroscopy.

English

&nbsp; In vitro&nbsp;and&nbsp;in vivo&nbsp;bio-stimulatory activity of a&nbsp;Lupinus albus&nbsp;cv. Bethuana white Seed Suspension (SS) was previously documented using vegetable crops. This was repeated for two grain crops, maize and wheat, in an attempt to provide a rationale forthe isolation and identification of the active compound(s) involved.&nbsp;In all cases, besides negative controls, the response to treatment with&nbsp;SS&nbsp;was compared to that of a commercial bio-stimulant, ComCat&reg;. At an optimum concentration of 5 mg â„“-1&nbsp;SS significantly enhanced seedling growth and yield in both crops. Subsequently, activity directed liquid-liquid extraction of ground&nbsp;L. albus&nbsp;seed resulted in an ethyl acetate extract containing most of the bio-stimulatory activity. Two&nbsp;in vitro&nbsp;bio-assay procedures were employed to appraise the&nbsp;in vitro&nbsp;bio-stimulatory properties of semi-purified fractions as well as pure compounds contained in it. The highly active ethyl acetate extract was fractionated further by means of adsorption column chromatography resulting in eighty five fractions that were combined into twelve on the basis of similarities in thin layer chromatography profiles. Four of these fractions showed significant bio-stimulatory activity and were pooled. From this one compound was isolated, purified and identified as glyceryl trilinoleate by means of nuclear magnetic resonance spectroscopy. &nbsp; Key words:&nbsp;Lupinus albus, seed suspension, field trials,&nbsp;yield, NMR spectroscopy, glyceryl trilinoleate.

Significant Modifications of the Salivary Proteome Potentially Associated with Complications of Down Syndrome Revealed by Top-down Proteomics

People with Down syndrome, a frequent genetic disorder in humans, have increased risk of health problems associated with this condition. One clinical feature of Down syndrome is the increased prevalence and severity of periodontal disease in comparison with the general population. Because saliva plays an important role in maintaining oral health, in the present study the salivary proteome of Down syndrome subjects was investigated to explore modifications with respect to healthy subjects. Whole saliva of 36 Down syndrome subjects, divided in the age groups 10-17 yr and 18-50 yr, was analyzed by a top-down proteomic approach, based on the high performance liquid chromatography-electrospray ionization-MS analysis of the intact proteins and peptides, and the qualitative and quantitative profiles were compared with sex- and age-matched control groups. The results showed the following interesting features: 1) as opposed to controls, in Down syndrome subjects the concentration of the major salivary proteins of gland origin did not increase with age; as a consequence concentration of acidic proline rich proteins and S cystatins were found significantly reduced in older Down syndrome subjects with respect to matched controls; 2) levels of the antimicrobial α-defensins 1 and 2 and histatins 3 and 5 were significantly increased in whole saliva of older Down syndrome subjects with respect to controls; 3) S100A7, S100A8, and S100A12 levels were significantly increased in whole saliva of Down syndrome subjects in comparison with controls. The increased level of S100A7 and S100A12 may be of particular interest as a biomarker of early onset Alzheimer's disease, which is frequently associated with Down syndrome.

Involvement of bradykinin and prostaglandins in the diuretic effects of Achillea millefolium L. (Asteraceae)

Ethnopharmacological relevanceAchillea millefolium L. (Asteraceae), popularly known as “mil-folhas”, is well recognized and widely used in Brazilian folk medicine to treat heart and kidney disorders. Among its popularly described effects are diuretic and hypotensive actions. Aim of the studyThe diuretic activity of Achillea millefolium L. extracts and its semi-purified fractions, as well as the mechanisms involved, were evaluated in male Wistar rats. Material and methodsAn aqueous extract (AEAM, 125–500mg/kg), hydroethanolic extract (HEAM, 30–300mg/kg), dichloromethane subfractions (DCM-2, 10 and 30mg/kg), or hydrochlorothiazide (10mg/kg), were orally administered and the animals were kept in metabolic cages for 8h for urine collection. To evaluate the involvement of bradykinin and prostaglandins in the diuretic action of Achillea millefolium, selected groups of rats received HOE-140 (1.5mg/kg, i.p.) or indomethacin (5mg/kg, p.o.), before treatment with a DCM-2 subfraction (30mg/kg). The urinary volume, conductivity, pH, density and electrolyte excretion were measured. ResultsSimilar to hydrochlorothiazide, both HEAM and DCM-2, but not AEAM, increased urinary volume and the excretion of Na+ and K+ when compared with the control group (vehicle). The diuretic effect of DCM-2 was abolished by HOE-140 (a bradykinin B2 receptor antagonist), as well as by indomethacin (a cyclooxygenase inhibitor). ConclusionThe present study reveals that extracts obtained from Achillea millefolium are able to effectively increase diuresis when orally administered in rats. This effect depends on both the activation of bradykinin B2 receptors and the activity of cyclooxygenases.

Metabolomic-Based Study of the Leafy Gall, the Ecological Niche of the Phytopathogen Rhodococcus fascians, as a Potential Source of Bioactive Compounds

Leafy gall is a plant hyperplasia induced upon Rhodococcus fascians infection. Previously, by genomic and transcriptomic analysis, it has been reported that, at the early stage of symptom development, both primary and secondary metabolisms are modified. The present study is based on the hypothesis that fully developed leafy gall, could represent a potential source of new bioactive compounds. Therefore, non-targeted metabolomic analysis of aqueous and chloroform extracts of leafy gall and non-infected tobacco was carried out by 1H-NMR coupled to principal component analysis (PCA) and orthogonal projections to latent structures-discriminant analysis (OPLS-DA). Polar metabolite profiling reflects modifications mainly in the primary metabolites and in some polyphenolics. In contrast, main modifications occurring in non-polar metabolites concern secondary metabolites, and gas chromatography and mass spectrometry (GC-MS) evidenced alterations in diterpenoids family. Analysis of crude extracts of leafy galls and non-infected tobacco leaves exhibited a distinct antiproliferative activity against all four tested human cancer cell lines. A bio-guided fractionation of chloroformic crude extract yield to semi-purified fractions, which inhibited proliferation of glioblastoma U373 cells with IC50 between 14.0 and 2.4 μg/mL. Discussion is focused on the consequence of these metabolic changes, with respect to plant defense mechanisms following infection. Considering the promising role of diterpenoid family as bioactive compounds, leafy gall may rather be a propitious source for drug discovery.

New insights on arthropod toxins that potentiate erectile function

The use of natural substances for the treatment of diseases or injuries is an ancient practice of many cultures. According to folklore, natural aphrodisiacs may help to raise libido and increase desire. The supposed aphrodisiacs mainly include a plethora of preparations of plants, among other substances. However, the real boundary between myth and reality has not been established yet in most cases and such boundaries must be drawn by scientific methods. A growing interest of the scientific community has been focused on animal venoms, especially those from arthropods, i.e. spiders and scorpions, which cause priapism, a prolonged and painful erection. This review highlights the studies that have been performed with venoms and toxins from arthropods known to cause priapism, among other toxic symptoms, pointing out some pharmacological approaches for better understanding this effect. To date, the venom of some spiders, mainly Phoneutria nigriventer, and scorpions, such as the yellow South American scorpion Tityus serrulatus, among others, have been known to cause priapism. Since erectile dysfunction (ED) is a growing health problem in the world, more common in patients with vascular diseases as diabetes and hypertension, the use of animal venoms and toxins as pharmacological tools could not only shed light to the mechanisms involved in erectile function, but also represent a possible model for new drugs to treat ED. Unfortunately, attempts to correlate the structure of those priapism-related toxins were unfruitful. Such difficulties lie firstly on the poor data concerning purified priapism-related toxins, instead of whole venoms and/or semi-purified fractions, and secondly, on the scarce available primary sequences and structural data, mainly from spider toxins. It has been shown that all these toxins modify the sodium (Na+) channel activity, mostly slowing down its inactivation current. Improving the knowledge on the tertiary structure of these toxins could provide a key in the search of a new drug for ED treatment.