Studies on the antimicrobial activity and brine shrimp toxicity of Zeyheria tuberculosa (Vell.) Bur. (Bignoniaceae) extracts and their main constituents

<p>Abstract</p> <p>Background</p> <p>Due to the indiscriminate use of antimicrobial drugs, the emergence of human pathogenic microorganisms resistant to major classes of antibiotics has been increased and has caused many clinical problems in the treatment of infectious diseases. Thus, the aim of this study was to evaluate for the first time the <it>in vitro </it>antimicrobial activity and brine shrimp lethality of extracts and isolated compounds from <it>Zeyheria tuberculosa </it>(Vell.) Bur., a species used in Brazilian folk medicine for treatment of cancer and skin diseases.</p> <p>Methods</p> <p>Using the disc diffusion method, bioautography assay and brine shrimp toxicity test (<it>Artemia salina </it>Leach), we studied the antimicrobial activity and lethality of extracts and isolated compounds against three microorganisms strains, including Gram-positive (<it>Staphylococcus aureus</it>) and Gram-negative (<it>Pseudomonas aeruginosa</it>) bacteria and yeasts (<it>Candida albicans</it>).</p> <p>Results</p> <p>In this study, the extracts inhibited <it>S. aureus </it>(8.0 ± 0.0 to 14.0 ± 0.0 mm) and <it>C. albicans </it>(15.3 ± 0.68 to 25.6 ± 0.4 mm) growth. In the brine shrimp test, only two of them showed toxic effects (LC₅₀29.55 to 398.05 μg/mL) and some extracts were non-toxic or showed weak lethality (LC₅₀705.02 to > 1000 μg/mL). From these extracts, four flavones [5,6,7,8-tetramethoxyflavone (1), 5,6,7-trimethoxyflavone (2), 4'-hydroxy-5,6,7,8-tetramethoxyflavone (3), and 4'-hydroxy-5,6,7-trimethoxyflavone (4)] were isolated through bioassay-guided fractionation and identified based on the 1D and 2D NMR spectral data. By bioautography assays, compounds 1 [<it>S. aureus </it>(16.0 ± 0.0 mm) and <it>C. albicans </it>(20.0 ± 0.0 mm)] and 3 [<it>S. aureus </it>(10.3 ± 0.6 mm) and <it>C. albicans </it>(19.7 ± 0.6 mm)] inhibited both microorganisms while 2 inhibited only <it>S. aureus </it>(11.7 ± 0.6 mm). Compound 4 did not restrain the growth of any tested microorganism.</p> <p>Conclusion</p> <p>Our results showed that extracts and isolated flavones from <it>Z. tuberculosa </it>may be particularly useful against two pathogenic microorganisms, <it>S. aureus </it>and <it>C. albicans</it>. These results may justify the popular use this species since some fractions tested had antimicrobial activity and others showed significant toxic effects on brine shrimps. However, in order to evaluate possible clinical application in therapy of infectious diseases, further studies about the safety and toxicity of isolated compounds are needed.</p

Anti-hepatocellular carcinoma activity using human HepG2 cells and hepatotoxicity of 6-substituted methyl 3-aminothieno[3,2-b]pyridine-2-carboxylate derivatives: in vitro evaluation, cell cycle analysis and QSAR studies

Hepatocellular carcinoma (HCC) is a highly complex cancer, resistant to commonly used treatments and new therapeutic agents are urgently needed. A total of thirty-two thieno[3,2-b]pyridine derivatives of two series: methyl 3-amino-6-(hetero)arylthieno[3,2-b]pyridine-2-carboxylates (1a-1t) and methyl 3-amino-6-[(hetero)arylethynyl]thieno[3,2-b]pyridine-2-carboxylates (2a-2n), previously prepared by some of us, were evaluated as new potential anti-HCC agents by studying their in vitro cell growth inhibition on human HepG2 cells and hepatotoxicity using a porcine liver primary cell culture (PLP1). The presence of amino groups linked to a benzene moiety emerges as the key element for the anti-HCC activity. The methyl 3-amino-6-[(3-aminophenyl)ethynyl]thieno[3,2-b]pyridine-2-carboxylate (2f) is the most potent compound presenting GI50 values on HepG2 cells of 1.2 μM compared to 2.9 μM of the positive control ellipticine, with no observed hepatotoxicity (PLP1 GI50>125 μM against 3.3 μM of ellipticine). Moreover this compound changes the cell cycle profile of the HepG2 cells, causing a decrease in the % of cells in the S phase and a cell cycle arrest in the G2/M phase. QSAR studies were also performed and the correlations obtained using molecular and 1D descriptors revealed the importance of the presence of amino groups and hydrogen bond donors for anti-HCC activity, and hydrogen bond acceptors for hepatotoxicity. The best correlations were obtained with 3D descriptors belonging to different subcategories for anti-HCC activity and hepatotoxicity, respectively. These results point to different molecular mechanisms of action of the compounds in anti-HCC activity and hepatotoxicity. This work presents some promising thieno[3,2-b]pyridine derivatives for potential use in the therapy of HCC. These compounds can also be used as scaffolds for further synthesis of more potent analogues.FCT, FEDER/COMPETE/QREN/E

Evaluation of the antioxidant activity of extracts obtained form cherry seeds.

Annual cherry production in Portugal is around 19,000 tonnes, in an area of about 6,450 ha and covering about 11,100 farms, concentrated in some northern and central interior territories. It is also in these regions that in recent decades there has been a significant increase in farms specialized in the production of cherry, using new cultivars and new technologies in a business production model. Apart from being consumed in fresh form, cherries are used for many food preparations, like sweets, jellies or confectionary. In the plants that transform cherries, a significant amount of cherry seeds (also called cherry pits) is generated as residue or waste. The possible usage of these residues as raw material for extraction of compounds with antioxidant properties is beneficial in term of economic value as well as environmental impact. Hence, the objective of this work was to obtain extract rich in compounds with antioxidant activity from cherry seeds. The cherry seeds were obtained from a local waste management company, Nutrofertil, located in Tondela, in the district of Viseu (Portugal). They were grinded and then submitted to extraction procedures testing different operating conditions: magnetic stirrer versus ultrasound, different solvents (methanol, ethanol, water) and temperatures (from 35 ºC to 80 ºC). For the obtained extracts antioxidant activity was evaluated through spectrophotometric methods, using the DPPH (2,2-diphenyl-1-picrylhydrazyl) and ABTS (2,2′-azino-di-(3-ethylbenzthiazoline sulfonic acid)) radicals, and also the Ferric Reducing Antioxidant Power Assay (FRAP). All measurements were replicated at least trice, and were expressed as mg Trolox equivalents per gram (mg TE/g). The results obtained for the different experimental conditions indicated that least efficient extractions at ambient temperature were obtained with methanol using magnetic stirrer and with water using ultrasounds, for which the antioxidant activities measured by the DPPH method were 0.26 and 0.33 mg TE/g and by the ABTS method were 0.82 and 0.86 mg TE/g, respectively. Most efficient methods were water:ethanol (at 50% concentration) and water (100%), using magnetic stirrer in both cases. Highest antioxidant activity was obtained for water:ethanol by the DPPH method (0.72 mg TE/g) and for water (100%) by the ABTS method (1.25 mg TE/g). Tests with different concentrations for the aqueous solutions of ethanol and at different temperatures revealed that with increasing concentration of water the antioxidant diminished, from 0.62 to 0.27 mg TE/g at 35 ºC using the DPPH method. Additionally, the variation in temperature allowed reaching a maximum extraction of compounds with antioxidant activity at 70 ºC and decreasing thereafter. The maximum values obtained were registered at 70 ºC for all cases and were 0.74 mg TE/g for the water:ethanol 50:50 (v/v) by the DPPH method, 2.16 mg TE/g for the water:ethanol 60:40 (v/v) by the ABTS method and 3.43 mg TE/g for the water:ethanol 60:40 (v/v) by the FRAP method. The results obtained by the different methods were concordant in terms of the observed trends but giving different values of the measured antioxidant activity, which is a common characteristic observed in these types of evaluation techniques. This research allowed establishing some operational conditions that should be selected in order to maximize the extraction of compounds with antioxidant activity from cherry seeds. The use of ultrasounds was not found beneficial and the magnetic stirrer technique revealed to be more useful. Also the use of methanol was not found suitable, which is a good point given that this solvent is more pollutant and has more problems of toxicity. With respect to temperature, it was found that temperatures higher than 70 ºC are not beneficial because they induce the degradation of some bioactive compounds thus reducing the antioxidant activity of thee extracts.info:eu-repo/semantics/publishedVersio

Aminodi(hetero)arylamines in the thieno[3,2-b]pyridine series: synthesis, effects in human tumor cells growth, cell cycle analysis, apoptosis and evaluation of toxicity using non-tumor cells

Three aminodi(hetero)arylamines were prepared via a palladium-catalyzed C-N Buchwald-Hartwig coupling of methyl 3-aminothieno[3,2-b]pyridine-2-carboxylate with different bromonitrobenzenes, followed by reduction of the nitro groups of the coupling products to the corresponding amino compounds. The aminodi(hetero)arylamines thus obtained were evaluated for their growth inhibitory effect on four human tumor cell lines MCF-7 (breast adenocarcinoma), A375-C5 (melanoma), NCI-H460 (non-small cell lung cancer) and HepG2 (hepatocellular carcinoma). The toxicity to non-tumor cells was also evaluated using a porcine liver primary cell culture (PLP1), established by us. The aminodi(hetero)arylamine with the NH2 group in the ortho position and an OMe group in the para position to the NH of the di(hetero)arylamine, is the most promising compound giving the lowest GI50 values (1.30–1.63 μM) in all the tested human tumor cell lines, presenting no toxicity to PLP1 at those concentrations. The effect of this compound on the cell cycle and induction of apoptosis was analyzed in the NCI-H460 cell line. It was observed that it altered the cell cycle profile causing a decrease in the percentage of cells in the G0/G1 phase and an increase of the apoptosis levels.Foundation for the Science and Technology (FCT–Portugal) for financial support through the NMR Portuguese network (Bruker 400 Avance III-Univ Minho). FCT and FEDER (European Fund for Regional Development) for financial support through the research centers PEst-C/QUI/UI686/2011and PEst-OE/AGR/UI0690/2011, the research project PTDC/QUI-QUI/111060/2009 and the post-Doctoralgrants attributed to R.C.C. and R.T.L. (SFRH/BPD/68344/2010 and SFRH/BPD/68787/2010, respectively). IPATIMUP is an Associate Laboratory of the Portuguese Ministry of Science, Technology and Higher Education and is partially supported by FCT

New di(hetero)arylethers and di(hetero)arylamines in the thieno[3,2-b]pyridine series: Synthesis, growth inhibitory activity on human tumor cell lines and non-tumor cells, effects on cell cycle and on programmed cell death

New fluorinated and methoxylated di(hetero)arylethers and di(hetero)arylamines were prepared functionalizing the 7-position of the thieno[3,2-blpyridine, using copper (C-O) or palladium (C N) catalyzed couplings, respectively, of the 7-bromothieno[3,2-blpyridine, also prepared, with ortho, meta and para fluoro or methoxy phenols and anilines. The compounds obtained were evaluated for their growth inhibitory activity on the human tumor cell lines MCF-7 (breast adenocarcinoma), NCI-H460 (non-small cell lung cancer), HCI15 (colon carcinoma), HepG2 (hepatocellular carcinoma) and HeLa (cervical carcinoma). The most active compounds, a di(hetero)arylether with a methoxy group in the meta position relative to the ether function and two di(hetero)arylamines with a methoxy group either in the ortho or in the meta position relative to the NH, were further tested at their GI(50) concentrations on NCI-H460 cells causing pronounced alterations in the cell cycle profile and a strong and significant increase in the programmed death of these cells. The fluorinated and the other methoxylated compounds did not show important activity, presenting high GI(50) values in all the cell lines tested. Furthermore, the hepatotoxicity of the compounds was assessed using porcine liver primary cells (PLP2), established by some of us. Results showed that one of the most active compounds was not toxic to the non-tumor cells at their GI(50) concentrations showing to be the most promising as antitumoral.The authors would like to thank to the Foundation for the Science and Technology (PCT Portugal) for financial support through the NMR Portuguese network (Bruker 400 Avance III-Univ Minho); to FCT and FEDER-COMPETE/QREN/EU for financial support through the research unities PEst-C/QUI/UI686/2011 and PEst-OE/AGR/UI0690/2011, the research project PTDC/QUI-QUI/111060/2009 and the post-Doctoral grants attributed to R.C.C. (SFRH/BPD/68344/2010) and R.T.L. (SRH/BPD/68787/2010). IPATIMUP is an Associate Laboratory of the Portuguese Ministry of Science, Technology and Higher Education and is partially supported by FCT

Genome-wide diversity and differentiation in New World populations of the human malaria parasite Plasmodium vivax.

BACKGROUND: The Americas were the last continent colonized by humans carrying malaria parasites. Plasmodium falciparum from the New World shows very little genetic diversity and greater linkage disequilibrium, compared with its African counterparts, and is clearly subdivided into local, highly divergent populations. However, limited available data have revealed extensive genetic diversity in American populations of another major human malaria parasite, P. vivax. METHODS: We used an improved sample preparation strategy and next-generation sequencing to characterize 9 high-quality P. vivax genome sequences from northwestern Brazil. These new data were compared with publicly available sequences from recently sampled clinical P. vivax isolates from Brazil (BRA, total n = 11 sequences), Peru (PER, n = 23), Colombia (COL, n = 31), and Mexico (MEX, n = 19). PRINCIPAL FINDINGS/CONCLUSIONS: We found that New World populations of P. vivax are as diverse (nucleotide diversity π between 5.2 × 10-4 and 6.2 × 10-4) as P. vivax populations from Southeast Asia, where malaria transmission is substantially more intense. They display several non-synonymous nucleotide substitutions (some of them previously undescribed) in genes known or suspected to be involved in antimalarial drug resistance, such as dhfr, dhps, mdr1, mrp1, and mrp-2, but not in the chloroquine resistance transporter ortholog (crt-o) gene. Moreover, P. vivax in the Americas is much less geographically substructured than local P. falciparum populations, with relatively little between-population genome-wide differentiation (pairwise FST values ranging between 0.025 and 0.092). Finally, P. vivax populations show a rapid decline in linkage disequilibrium with increasing distance between pairs of polymorphic sites, consistent with very frequent outcrossing. We hypothesize that the high diversity of present-day P. vivax lineages in the Americas originated from successive migratory waves and subsequent admixture between parasite lineages from geographically diverse sites. Further genome-wide analyses are required to test the demographic scenario suggested by our data

Epidemiology of Disappearing Plasmodium vivax Malaria: A Case Study in Rural Amazonia

Background: New frontier settlements across the Amazon Basin pose a major challenge for malaria elimination in Brazil. Here we describe the epidemiology of malaria during the early phases of occupation of farming settlements in Remansinho area, Brazilian Amazonia. We examine the relative contribution of low-density and asymptomatic parasitemias to the overall Plasmodium vivax burden over a period of declining transmission and discuss potential hurdles for malaria elimination in Remansinho and similar settings. Methods: Eight community-wide cross-sectional surveys, involving 584 subjects, were carried out in Remansinho over 3 years and complemented by active and passive surveillance of febrile illnesses between the surveys. We used quantitative PCR to detect low-density asexual parasitemias and gametocytemias missed by conventional microscopy. Mixed-effects multiple logistic regression models were used to characterize independent risk factors for P. vivax infection and disease. Principal Findings/Conclusions P. vivax prevalence decreased from 23.8% (March–April 2010) to 3.0% (April–May 2013), with no P. falciparum infections diagnosed after March–April 2011. Although migrants from malaria-free areas were at increased risk of malaria, their odds of having P. vivax infection and disease decreased by 2–3% with each year of residence in Amazonia. Several findings indicate that low-density and asymptomatic P. vivax parasitemias may complicate residual malaria elimination in Remansinho: (a) the proportion of subpatent infections (i.e. missed by microscopy) increased from 43.8% to 73.1% as P. vivax transmission declined; (b) most (56.6%) P. vivax infections were asymptomatic and 32.8% of them were both subpatent and asymptomatic; (c) asymptomatic parasite carriers accounted for 54.4% of the total P. vivax biomass in the host population; (d) over 90% subpatent and asymptomatic P. vivax had PCR-detectable gametocytemias; and (e) few (17.0%) asymptomatic and subpatent P. vivax infections that were left untreated progressed to clinical disease over 6 weeks of follow-up and became detectable by routine malaria surveillance

Purification of a lectin from Cratylia mollis crude extract seed by a single step PEG/phosphate aqueous two-phase system

The partitioning and purification of lectins from the crude extract of Cratylia mollis seeds (Cramoll 1,4) was investigated in aqueous two-phase systems (ATPS). A factorial design model (24) was used to evaluate the influence of polyethylene glycol (PEG) molar mass (15008000g/mol), PEG concentration (12.517.5% w/w), phosphate (1015% w/w) concentration, and pH (68) on the differential partitioning, purification factor, and yield of the lectin. Polymer and salt concentration were the most important variables affecting partition of lectin and used to find optimum purification factor by experimental BoxBehnken design together with the response surface methodology (RSM). ATPS showed best conditions composed by 13.9% PEG1500, 15.3% phosphate buffer at pH 6, which ensured purification factor of 4.70. Sodium dodecyl sulfatepolyacrylamide gel electrophoresis showed a single band of protein with 26.1kDa. Furthermore, results demonstrated a thermostable lectin presenting activity until 60°C and lost hemagglutinating activity at 80°C. According to the obtained data it can be inferred that the ATPS optimization using RSM approach can be applied for recovery and purification of lectins.We are grateful to the following bodies for the grants awarded: CAPES (Coordination for the Improvement of Level Personnel Superior); FACEPE (Pernambuco Science and Technology Foundation): Researcher's scholarship grant: BFP-0017-5.05/18 CNPq (National Council for Scientific Development and Technological) process: 427153/2016-6 and we also thank the reviewers for their valuable comments and suggestions as these helped us to improve the manuscript.info:eu-repo/semantics/publishedVersio