Antibacterial activity of extracts and neolignans from Piper regnellii (Miq.) C. DC. var. pallescens (C. DC.) Yunck

The evaluation of the activity of the aqueous and ethyl acetate extracts of the leaves of Piper regnellii was tested against gram-positive and gram-negative bacteria. The aqueous extract displayed a weak activity against Staphylococcus aureus and Bacillus subtilis with minimal inhibitory concentration (MIC) and minimal bactericidal concentration (MBC) of 1000 micrograms/ml. The ethyl acetate extract presented a good activity against S. aureus and B. subtilis with MIC and MBC at 15.62 micrograms/ml. In contrast to the relative low MICs for gram-positive bacteria, gram-negative bacteria were not inhibited by the extracts at concentrations < or = 1000 mg/ml. The ethyl acetate extract was fractionated on silica gel into nine fractions. The hexane and chloroform fractions were active against S. aureus (MIC at 3.9 micrograms/ml) and B. subtilis (MIC at 3.9 and 7.8 micrograms/ml, respectively). Using bioactivity-directed fractionation, the hexane fraction was rechromatographed to yield the antimicrobial compounds 1, 2, 5, and 6 identified as eupomatenoid-6, eupomatenoid-5, eupomatenoid-3, and conocarpan, respectively. The pure compounds 1 and 2 showed a good activity against S. aureus with MIC of 1.56 micrograms/ml and 3.12 micrograms/ml, respectively. Both compounds presented MIC of 3.12 micrograms/ml against B. subtilis. The pure compound 6 named as conocarpan was quite active against S. aureus and B. subtilis with MIC of 6.25 micrograms/ml. The antibacterial properties of P. regnellii justify its use in traditional medicine for the treatment of wounds, contaminated through bacteria infections.     

Anti-Helicobacter pylori activity of the methanolic extract of Geum iranicum and its main compounds

Geum iranicum Khatamsaz, belonging to the Rosaceae family, is an endemic plant of Iran. The methanol extract of the roots of this plant showed significant activity against one of the clinical isolates of Helicobacter pylori which was resistant to metronidazole. The aim of this study was the isolation and evaluation of the major compounds of G. iranicum effective against H. pylori. The compounds were isolated using various chromatographic methods and identified by spectroscopic data (1H and 13C NMR, HMQC, HMBC, EI-MS). An antimicrobial susceptibility test was performed employing the disk diffusion method against clinical isolates of H. pylori and a micro dilution method against several Gram-positive and Gram-negative bacteria; additionally the inhibition zone diameters (IZD) and minimum inhibitory concentrations (MIC) values were recorded. Nine compounds were isolated: two triterpenoids, uvaol and niga-ichigoside F1, three sterols, beta-sitosterol, beta-sitosteryl acetate, and beta-sitosteryl linoleate, one phenyl propanoid, eugenol, one phenolic glycoside, gein, one flavanol, (+)-catechin, and sucrose. The aqueous fraction, obtained by partitioning the MeOH extract with water and chloroform, was the most effective fraction of the extract against all clinical isolates of H. pylori. Further investigation of the isolated compounds showed that eugenol was effective against H. pylori but gein, diglycosidic eugenol, did not exhibit any activity against H. pylori. The subfraction D4 was the effective fraction which contained tannins. It appeared that tannins were probably the active compounds responsible for the anti-H. pylori activity of G. iranicum. The aqueous fraction showed a moderate inhibitory activity against both Gram-positive and Gram-negative bacteria. The MIC values indicated that Gram-positive bacteria including Staphylococcus aureus, Staphylococcus epidermidis, and Bacillus subtilis are more susceptible than Gram-neagative bacteria including Escherichia coli and Pseudomonas aeruginosa.     

Antibacterial activity of extract, fractions and four compounds extracted from Piper solmsianum C. DC. VAR. solmsianum (Piperaceae)

Piper solmsianum C. DC. var. solmsianum (Piperaceae) is a shrub commonly found in areas with wet tropical soils. Other Piper species have been used in folk medicine as antitumoral and antiseptic agents. We studied the crude methanolic extract, some organic fractions and compounds isolated from this plant for possible antimicrobial activity against Gram-positive and Gram-negative bacteria. The bioautographic assays disclosed three inhibition zones. The minimal inhibitory concentration (MIC) and minimal bactericidal concentration (MBC) were determined showing excellent activity, particularly against the Gram-positive bacteria (Bacillus cereus, Staphylococcus aureus, Staphylococcus saprophyticus and Streptococcus agalactiae). It appears that the antimicrobial activity of Piper solmsianum is related mainly to the presence of conocarpan and eupomatenoid-5 (neolignans). However another, as yet unidentified, active compound could also be extracted from the plant.     

Two fractions required for cell-free protein synthesis with components from rabbit reticulocytes

An extract was prepared from rabbit reticulocyte ribosomes after treatment with potassium chloride as described by Miller, Hamada, Yang, Cohen & Schweet (1967). This extract has been shown to convert monoribosomes into polyribosomes during protein synthesis in vitro (Cohen, 1968). The nature of this extract was studied in greater detail. Centrifugation of the extract through a sucrose density gradient separated the activity into a fast-sedimenting fraction. The two fractions were shown to have different functions in stimulating cell-free protein synthesis and their active components were shown to be protein or partly protein in nature. Each fraction was analysed by electrophoresis and in the analytical ultracentrifuge. It was concluded that the active component in the fast-sedimenting fraction had a sedimentation coefficient of 15.5s and that of the slow-sedimenting fraction 10.5s.     

<Emphasis Type="Italic">Lyso</Emphasis>-phosphatidylethanolamine-enhanced phenylalanine ammonia-lyase and insoluble acid invertase in isolated radish cotyledons

Application of lyso-phosphatidylethanolamine (LPE) is purported to suppress fruit ripening and delay foliar senescence. However, the endogenous LPE response of plants is more typically associated with propagation of wound and stress signals. Experiments were therefore carried out to determine whether exogenous LPE could elicit defense responses in plants by determining the effect of this lyso-phospholipid on activity of two key metabolic enzymes and pathogenesis-related proteins viz phenylalanine ammonia-lyase (PAL; EC 4.3.1.5) and insoluble acid invertase (Ac INV; EC 3.2.1.26) in expanding cotyledons of Raphanus sativus L. cv. Cherry Belle (radish). Activity of both enzymes was increased following exposure of tissue to 18:0-LPE and the response was dose dependent. Soluble Ac INV activity was not enhanced by exogenous 18:0-LPE. Increased PAL activity appeared to coincide with a decline in phenolic acid content and a rise in sinapine and lignin. An increase in insoluble Ac INV by 18:0-LPE was associated with a reduction in sucrose concentration. However, levels of glucose and fructose were unaffected. In view of these findings it is proposed that applied LPE acts to co-ordinate carbohydrate partitioning locally to fulfill anabolic respiratory requirements associated with the propagation of systemic wound and stress responses. Furthermore, the impact of exogenous 18:0-LPE on insoluble Ac INV activity is discussed in relation to the proposed role of this enzyme in cytokinin-mediated senescence delay.     

Antimicrobial activity of leaf extract of Basilicum polystachyon (L) Moench

Phenolic extract of leaves of Basilicum polystachyon (L) Moench was tested for in vitro antimicrobial activity against five bacteria (Escherichia coli, Pseudomonas aeruginosa, Staphylococcus aureus, Bacillus subtilis, Micrococcus leuteus) and three fungi (Fusarium oxysporum, Aspergillus niger, Helminthosporium oryzae). Efficacy of organic solvents, methanol and ethanol, as agents for extraction was compared with acidic water (2M; HCl). High-pressure liquid chromatographic (HPLC) data showed that acidic extraction (2M; HCl) resulted in higher yield of caffeic acid (0.437 mg g(-1)) and rosmarinic acid (0.919 mg g(-1)). Acidic extract showed high activity against Gram (+) ve bacteria, but was less active against Gram (-) ve bacteria. Amongst the tested fungi, maximum activity was exhibited against Aspergillus niger. This is the first report on the phenolic constituents and bioactivity of B. polystachyon.     

Antiviral activity of Rwandan medicinal plants against human immunodeficiency virus type-1 (HIV-1)

Selected plants used in Rwandan traditional medicine for the treatment of infections and/or rheumatoid diseases were investigated for antiviral activity in vitro against human immunodeficiency virus type-1 (HIV-1). Of the 38 tested 80% ethanolic extracts, belonging to plants of 21 different families only the extracts from the leaves of Aspilia pluriseta (Asteraceae) and Rumex bequaertii (Polygonaceae) had interesting selectivity indices (SI = ratio of the 50% cytotoxic concentration to the 50% effective antiviral concentration) higher than 1. Further fractionation of the initially antivirally inactive ethanolic extract of Tithonia diversifolia, however, led to an aqueous fraction with a high anti-HIV-1 activity (SI > 461), indicating that the cytotoxicity of some plant components may mask the antiviral properties of the active plant substances in total plant extracts.     

Inhibition of lipopolysaccharid-induced sickness behavior by a dry extract from the roots of Pelargonium sidoides (EPs® 7630) in mice

The host response to infections comprise the synthesis and release of proinflammatory cytokines (e.g. IL-1ß, TNF-α, IL-6) which induce symptoms of sickness behavior characterised by anorexia, depressed activity, listlessness or malaise. In laboratory animals, sickness behavior can be induced by the administration of cytokines itself or by cytokine-inducers such as lipopolysaccharide (LPS), the active fragment of endotoxin from Gram-negative bacteria.Preparations from roots of Pelargonium sidoides have been traditionally used in South African folk medicine for the treatment of different diseases (e.g. diarrhea, dysmenorrhea, hepatic disorders and respiratory tract infections including tuberculosis). Today, aqueous ethanolic extracts of Pelargonium sidoides are marketed mainly for respiratory tract infections. We studied the effects of the extract EPs^® 7630 and different fractions separated by ultrafiltration in an animal model of sickness behavior.The results of this study demonstrate that the extract EPs^® 7630 and the high-molecular weight fraction (F3) alleviate the symptoms of sickness behavior.     

Viper and cobra venom neutralization by β-sitosterol and stigmasterol isolated from the root extract of Pluchea indica Less. (Asteraceae)

We reported previously that the methanolic root extract of the Indian medicinal plant Pluchea indica Less. (Asteraceae) could neutralize viper venom-induced action [Alam, M.I., Auddy, B., Gomes, A., 1996. Viper venom neutralization by Indian medicinal plant (Hemidesmus indicus and P. indica) root extracts. Phytother. Res. 10, 58-61]. The present study reports the neutralization of viper and cobra venom by beta-sitosterol and stigmasterol isolated from the root extract of P. indica Less. (Asteraceae). The active fraction (containing the major compound beta-sitosterol and the minor compound stigmasterol) was isolated and purified by silica gel column chromatography and the structure was determined using spectroscopic analysis (EIMS, (1)H NMR, (13)C NMR). Anti-snake venom activity was studied in experimental animals. The active fraction was found to significantly neutralize viper venom-induced lethal, hemorrhagic, defibrinogenation, edema and PLA(2) activity. Cobra venom-induced lethality, cardiotoxicity, neurotoxicity, respiratory changes and PLA(2) activity were also antagonized by the active component. It potentiated commercial snake venom antiserum action against venom-induced lethality in male albino mice. The active fraction could antagonize venom-induced changes in lipid peroxidation and superoxide dismutase activity. This study suggests that beta-sitosterol and stigmasterol may play an important role, along with antiserum, in neutralizing snake venom-induced actions.     

Isolation, identification, and biological evaluation of Nrf2-ARE activator from the leaves of green perilla (Perilla frutescens var. crispa f. viridis)

The nuclear factor erythroid 2-related factor 2 (Nrf2)-antioxidant response element (ARE) pathway is a cellular defense system against oxidative stress. Activation of this pathway increases expression of antioxidant enzymes. Epidemiological studies have demonstrated that the consumption of fruits and vegetables is associated with reduced risk of contracting a variety of human diseases. The aim of this study is to find Nrf2-ARE activators in dietary fruits and vegetables. We first attempted to compare the potency of ARE activation in six fruit and six vegetables extracts. Green perilla (Perilla frutescens var. crispa f. viridis) extract exhibited high ARE activity. We isolated the active fraction from green perilla extract through bioactivity-guided fractionation. Based on nuclear magnetic resonance and mass spectrometric analysis, the active ingredient responsible for the ARE activity was identified as 2',3'-dihydroxy-4',6'-dimethoxychalcone (DDC). DDC induced the expression of antioxidant enzymes, such as γ-glutamylcysteine synthetase (γ-GCS), NAD(P)H: quinone oxidoreductase-1 (NQO1), and heme oxygenase-1. DDC inhibited the formation of intracellular reactive oxygen species and the cytotoxicity induced by 6-hydroxydopamine. Inhibition of the p38 mitogen-activated protein kinase pathway abolished ARE activation, the induction of γ-GCS and NQO1, and the cytoprotective effect brought about by DDC. Thus, this study demonstrated that DDC contained in green perilla enhanced cellular resistance to oxidative damage through activation of the Nrf2-ARE pathway.     

Delayed leaf senescence by exogenous lyso-phosphatidylethanolamine: Towards a mechanism of action

Exogenous application of the lysophospholipid, lyso-phosphatidylethanolamine (LPE) is purported to delay leaf senescence in plants. However, lyso-phospholipids are well known to possess detergent-like activity and application of LPE to plant tissues might be expected to rather elicit a wound-like response and enhance senescence progression. Since phosphatidic acid (PA) accumulation and leaf cell death are a consequence of wounding, PA- and hormone-induced senescence was studied in leaf discs from Philodendron cordatum (Vell.) Kunth plants in the presence or absence of egg-derived 18:0-LPE and senescence progression quantified by monitoring both lipid peroxidation (as the change in malondialdehyde concentration), and by measuring retention of total chlorophyll (Chl_a+b) and carotenoids (C_c+x). Only abscisic acid (ABA) stimulated lipid peroxidation whereas ABA, 1-aminocyclopropane-1-carboxylic acid (ACC), the immediate precursor to ethylene (ETH), and 16:0–18:2-PA stimulated loss of chloroplast pigments. Results using primary alcohols as attenuators of the endogenous PA signal confirmed a role for PA as an intermediate in both ABA- and ETH-mediated senescence progression. Exogenous 18:0-LPE did not appear to influence senescence progression and was unable to reverse hormone-induced senescence progression. However, when supplied together with 16:0–18:2-PA at 1:1 (mol:mol), activity of phosphatidylglycerol (PG) hydrolase, chlorophyllase (E.C. 3.1.1.14), and progression of leaf senescence were negated. This apparent anti-senescence activity of exogenous 18:0-LPE was associated with induction of the pathogenesis-related protein, extracellular acid invertase (Ac INV, E.C. 3.2.1.26) suggesting that 18:0-LPE like 16:0–18:2-PA functions as an elicitor.     

Isolation and identification of the probing stimulants in the rice plant for the white-back planthopper, Sogatella furcifera (homoptera: delphacidae)

Adult females of the white-back planthopper, Sogatella furcifera, showed characteristic behavior of stylet sheath deposit on a parafilm membrane when fed on a 2% aqueous crude rice leaf and stem extract containing 15% sucrose. Subsequent bioassays revealed that the butanol-soluble fraction of the extract was highly active against the insects. When the butanol fraction was chromatographed on an ODS open column and eluted in sequence with a mixture of an increasing concentration of methanol in water, the 40 % methanol fraction was separated as the most active. A further bioassay of the HPLC components in the active fraction revealed that two major components (1 and 3) stimulated the high probing activity of the white-back planthopper only when they were combined. Of the active components, one component (3) was identified to be tricin 5-O-glucoside by spectroscopic analyses.     

Anti-microfouling Activity of Lipidic Metabolites from the Invasive Brown Alga Sargassum muticum (Yendo) Fensholt

The purification of the chloroform extract from the brown invasive macroalga Sargassum muticum, through a series of chromatographic separations, yielded 12 fractions that were tested against strains of bacteria, microalgae, and fungi involved in marine biofilm formation. The chemical composition of four (a, c, g, and k) out of the six fractions that exhibited anti-microfouling activity was investigated. Fraction a contained saturated and unsaturated linear hydrocarbons (C₁₂–C₂₇). Arachidonic acid was identified as the major metabolite in fraction c whereas fraction g contained mainly palmitic, linolenic, and palmitoleic acids. Fraction k was submitted to further purification yielding the fraction kAcaF1e that was composed of galactoglycerolipids, active against the growth of two of the four bacterial strains (Shewanella putrefaciens and Polaribacter irgensii) and all tested fungi. These promising results, in particular the isolation and the activity of galactoglycerolipids, attest the potential of the huge biomass of S. muticum as a source of new environmentally friendly antifouling compounds.     

In vitro anti-Neisseria gonorrhoeae activity of Terminalia macroptera leaves

We used the agar dilution method to evaluate the antibacterial effect of Terminalia macroptera leaf (Tml) extract against nine reference and clinical Neisseria gonorrhoeae strains, including penicillin- and tetracycline-resistant and -susceptible strains. Tml possesses anti-N. gonorrhoeae activity against all of the strains and the minimum inhibitory concentrations (MIC) were between 100 and 200 microg ml(-1). We then used a liquid-liquid partition method to divide the Tml extract into five fractions and determined the anti-N. gonorrhoeae activity of each of the fractions. All of the fractions showed antibacterial activity. The most active one was identified as the diethyl ether fraction and had MIC values of between 25 and 50 microg ml(-1) against all of the strains.     

In vitro evaluation of Datura innoxia (thorn-apple) for potential antibacterial activity

Various parts of Datura innoxia were examined for potential antibacterial activity by preparing their crude aqueous and organic extracts against Gram-negative bacteria (Escherichia coli and Salmonella typhi) and Gram-positive bacteria (Bacillus cereus, Bacillus subtilis and Staphylococcus aureus). The results of agar well diffusion assay indicated that the pattern of inhibition depends largely upon the plant part, solvent used for extraction and the organism tested. Extracts prepared from leaves were shown to have better efficacy than stem and root extracts. Organic extracts provided potent antibacterial activity as compared to aqueous extracts. Among all the extracts, methanolic extract was found most active against almost all the bacterial species tested. Gram-positive bacteria were found most sensitive as compared to Gram-negative bacteria. Staphylococcus aureus was signifi cantly inhibited by almost all the extracts even at very low MIC followed by other Gram-positives. For Escherichia coli (a Gram-negative bacterium), the end point was not reached for ethyl acetate extract while it was very high for other extracts. The study promises an interesting future for designing a potentially active antibacterial agent from Datura innoxia.     

Effects of dentin proteins, transforming growth factor beta 1 (TGF beta 1) and bone morphogenetic protein 2 (BMP2) on the differentiation of odontoblast in vitro.

We have studied the effects of dentin proteins, of Transforming Growth Factor beta 1 (TGF beta 1) and Bone Morphogenetic Protein (BMP2) on the differentiation of odontoblasts in vitro. The total EDTA-soluble fraction of dentin proteins, prepared from rabbit incisors was further separated by chromatography on DEAE-Cellulose and heparin-agarose columns. While the total EDTA-soluble fraction of dentin had no effect on cultured dental papillae, fractions retained on both columns were able to initiate functional differentiation of preodontoblasts of isolated day-17 first lower mouse molar dental papillae cultured in vitro. TGF beta 1 and BMP2, both stimulated the matrix secretion by dental papillae cells. TGF beta 1 and BMP2, combined with the inactive total EDTA-soluble fraction, stimulated odontoblast differentiation. An active fraction retained on DEAE-Cellulose completely lost the inductive activity after incubation with a neutralizing anti-TGF beta antibody. These results demonstrate that a TGF beta-like molecule present in dentin could interact with some component which acts as a modulator of its activity on the initiation of the cytological and functional differentiation of odontoblasts.     

Pepper beta-galactosidase 1 (PBG1) plays a significant role in fruit ripening in bell pepper (Capsicum annuum)

During bell pepper (Capsicum annuum L.) fruit ripening, beta-galactosidase activity increased markedly as compared with other glycosidases. We purified 77.5 kDa exo-1,4-beta-D-galactanase from red bell pepper fruit classified as beta-galactosidase II. A marked decrease in galactose content appeared during fruit ripening, especially in the pectic fraction. The purified enzyme hydrolyzed a considerable amount of galactose residues in this fraction. We isolated bell pepper beta-galactosidase (PBG1) cDNA. This PBG1 protein contained the putative active site, G-G-P-[LIVM]-x-Q-x-E-N-E-[FY], belonging to glycosyl hydrolase family 35. Quantitative RT-PCR revealed that the expression of PBG1 in red fruit was significantly stronger than that from any other tissues. Moreover, expression of PBG1 occurred prior to that of pepper endo-polygalacturonase 1 (PPG1), the major fruit-ripening enzyme. Based on these results, it appears that the hydrolysis of galactose residues in pectic substances is the first event in the ripening process in bell pepper fruit.     

Antileishmanial activity of indole alkaloids from Aspidosperma ramiflorum

The present study was designated to evaluate the antileishmanial activity of acid and basic fractions that were obtained after acid-basic extraction, from ethanolic 70% crude extract and pure compounds from the stem bark of Aspidosperma ramiflorum. The basic alkaloidal fraction presented a good activity against the extracellular form (promastigotes) of Leishmania (L.) amazonensis (LD(50) value<47 microg/ml). Based on these findings, the basic fraction was fractionated on silica gel column chromatography in a bioassay-guided fractionation affording individual purified ramiflorines A and B. Both ramiflorines A and B showed significant activity against Leishmania (L.) amazonensis (LD(50) values of 16.3+/-1.6 microg/ml and 4.9+/-0.9 microg/ml, respectively). Our results are promising, showing that these compounds are biologically active against Gram-positive bacteria.     

Organic extracts with antimicrobian activity from Penicillium sp. (Moniliales) isolated from the sponge Ircinia felix (Porifera: Demospongiae)

This research expresses the potential of the bacterial activity present in the organic extracts obtained from Penicillium sp., isolated from the esponge Irciniafelix. This activity was evaluated through agar diffusion test and Minimal Inhibitory Concentration (MIC). The susceptibility trials of organic fractions were carried out against Staphylococcus aureus, S. epidermidis, Bacillus cereus and B. subtilis. The use of the chromatographic techniques (CLV and TLC), permitted to obtain bioactive organic extracts of different polarities, of which only the EtOAc and MeOH fractions inhibited the growth of the bacteria used. Of the EtOAc fractionation, only fraction number 3 EtOAc/Hex presented greatest activity against the Gram-positive bacteria. Number 1 EtOAc/Hex fraction increased its activity against S. aureus (24 mm) and S. epidermidis (25 mm), which can be explained by the loss of possible antagonistic effect during the fractionation process. The CMI trials were carried out for the EtOAc number I subfraction against S. aureus, S. epidermidis, B. cereus and B. subtilis, wich was clinical interest, and shows the potential of this organic extract as antimicrobial agent.     

Screening and characterization of microbial inhibitors against eukaryotic protein phosphatases (PP1 and PP2A)

AIM: To identify novel microbial inhibitors of protein phosphatase 1 (PP1). METHODS AND RESULTS: 750 actinomycetes and 408 microfungi were isolated from Sabah forest soils and screened for production of potential PP1 inhibitors using an in vivo screening system, in which candidate inhibitors were identified through mimicking the properties of PP1-deficient yeast cells. Acetone extracts of two fungi, H9318 (Penicillium) and H9978 (non-Penicillium) identified in this way showed inhibitory activity towards both mammalian PP1 and PP2A in an in vitro phosphatase assay, while extract from H7520 (Streptomyces) inhibited PP2A but not PP1. Consistently, using a drug-induced haploinsufficiency test, strains with either reduced PP1 or PP2A function were hypersensitive to H9318 and H9978 extracts whereas only the latter strain showed hypersensitivity to H7250 extract. H9318 extract was fractionated using RP-HPLC into two active peaks (S1 and S2). A yeast strain with reduced PP1 function showed hypersensitivity to fraction S2 whereas a strain with reduced PP2A function was hypersensitive to fraction S1. However, S1 and S2 inhibited both PP1 and PP2A activities to a similar extent. CONCLUSION: Three candidate PP inhibitors have been identified. SIGNIFICANCE AND IMPACT OF THE STUDY: Further development may generate useful research tools and ultimately therapeutic agents.