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1.
J Chromatogr A ; 1625: 461302, 2020 Aug 16.
Artigo em Inglês | MEDLINE | ID: mdl-32709345

RESUMO

Additives are added to polymers in small concentration to achieve desired application properties widely used to tailor the properties. The rapid diversification of their molecular structures, with often only minute differences, necessitates the development of adequate chromatographic techniques. While modified silica so far is the workhorse as stationary phase we have probed the potential of porous graphitic carbon (HypercarbTM) for this purpose. The results show that the multitude of physicochemical interactions between analyte molecules and the graphitic surface enables separations of polyolefin stabilizers with unprecedented selectivity. To support the chromatographic results the adsorption capability of HypercarbTM for selected antioxidants and UV absorbers has been determined by Raman spectroscopy and argon physisorption measurements. The shift of the Graphite-band in the Raman spectra of HypercarbTM upon infusion with additives correlates with the changes in the Adsorption Potential Distributions. The results of argon physisorption measurements go hand in hand with the chronology of desorption of the additives in liquid chromatography experiments. The elution sequence can be explained by van der Waals or London forces, π-π-interactions and electron lone pair donor-acceptor interactions between the graphite surface and analyte functional groups.


Assuntos
Grafite/química , Polímeros/química , Análise Espectral Raman , Adsorção , Antioxidantes/isolamento & purificação , Argônio/química , Clorofórmio/química , Éteres Metílicos/química , Polienos/química , Porosidade , Fatores de Tempo
2.
J Chromatogr A ; 1621: 461081, 2020 Jun 21.
Artigo em Inglês | MEDLINE | ID: mdl-32349863

RESUMO

The chemical composition distribution (CCD) of three single site made ethylene/1-octene copolymers was investigated through offline-hyphenation of solvent gradient interaction chromatography (SGIC) with 1H NMR. Thus, a clear, non-linear correlation between SGIC elution time and chemical composition was found under the specific measurement conditions applied here. The application of 1H NMR as detection allowed to determine the CCD with unprecedented accuracy. 2D-LC of the copolymers revealed the correlation between CCD and molar mass distribution (MMD) in a quantitative manner. Furthermore, this approach allowed a comparison between the response behavior of an evaporative light scattering detector (ELSD, semi-quantitative, commonly applied in SGIC) and that of an infrared (IR) detector (quantitative, commonly applied in SEC). As a result, it could be shown that ELSD results are close to IR results for the system investigated here, in other words, the often-criticized semi-quantitative response behavior of the ELSD is affecting results in an acceptable manner.


Assuntos
Alcenos/análise , Cromatografia Líquida de Alta Pressão/métodos , Elastômeros/análise , Etilenos/análise , Polienos/análise , Calibragem , Espectroscopia de Ressonância Magnética Nuclear de Carbono-13 , Elastômeros/química , Peso Molecular , Espectroscopia de Prótons por Ressonância Magnética , Solventes/química , Temperatura , Fatores de Tempo
3.
Nat Commun ; 11(1): 1502, 2020 03 20.
Artigo em Inglês | MEDLINE | ID: mdl-32198389

RESUMO

Although certain microbial lipids are toxins, the structural features important for cytotoxicity remain unknown. Increased functional understanding is essential for developing therapeutics against toxic microbial lipids. Group B Streptococci (GBS) are bacteria associated with preterm births, stillbirths, and severe infections in neonates and adults. GBS produce a pigmented, cytotoxic lipid, known as granadaene. Despite its importance to all manifestations of GBS disease, studies towards understanding granadaene's toxic activity are hindered by its instability and insolubility in purified form. Here, we report the synthesis and screening of lipid derivatives inspired by granadaene, which reveal features central to toxin function, namely the polyene chain length. Furthermore, we show that vaccination with a non-toxic synthetic analog confers the production of antibodies that inhibit granadaene-mediated hemolysis ex vivo and diminish GBS infection in vivo. This work provides unique structural and functional insight into granadaene and a strategy to mitigate GBS infection, which will be relevant to other toxic lipids encoded by human pathogens.


Assuntos
Hemólise , Lipídeos/química , Polienos/química , Nascimento Prematuro/microbiologia , Infecções Estreptocócicas/metabolismo , Adulto , Animais , Linfócitos B , Toxinas Bacterianas/química , Vacinas Bacterianas , Linfócitos T CD4-Positivos , Modelos Animais de Doenças , Feminino , Humanos , Recém-Nascido , Lipídeos/imunologia , Lipídeos/toxicidade , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Polienos/imunologia , Gravidez , Infecções Estreptocócicas/imunologia , Infecções Estreptocócicas/microbiologia , Infecções Estreptocócicas/prevenção & controle , Streptococcus agalactiae , Vacinação
4.
J Biosci Bioeng ; 129(5): 558-564, 2020 May.
Artigo em Inglês | MEDLINE | ID: mdl-31924510

RESUMO

ε-Poly-l-lysine (ε-PL) produced as a secondary metabolite of Streptomyces albulus has long been used as a natural food preservative in a number of countries, including Japan, the United States, South Korea, and China. To date, numerous studies employing classical biotechnological approaches have been carried out to improve its productivity. Here we report a modern and rational genetic approach to enhancing metabolic flux toward ε-PL biosynthesis. Based on in silico genome analyses, we revealed that S. albulus NBRC14147 produces five antifungal polyene antibiotics-tetramycin A and B, tetrin A and B, and a trace amount of nystatin A1-concomitantly with antimicrobial ε-PL. Targeted inactivation of the biosynthetic gene cluster for tetramycins and tetrins in a nystatin A1 production-deficient mutant completely abolished the production of polyene macrolides, which in turn led to an approximately 20% improvement in ε-PL production that closely correlated with the polyene defects. The biosynthetic flux for ε-PL was thus successfully enhanced by inactivation of the concomitant secondary metabolite biosynthetic pathways. Since this elimination of concomitantly produced metabolites also allows for simpler purification after fermentation production of ε-PL, the rational strain engineering strategy we show here will improve its industrial production.


Assuntos
Macrolídeos/metabolismo , Polienos/metabolismo , Polilisina/biossíntese , Streptomyces/metabolismo , Fermentação , Conservantes de Alimentos/metabolismo , Macrolídeos/química , Polienos/química , Streptomyces/química , Streptomyces/genética
5.
Microb Cell Fact ; 19(1): 3, 2020 Jan 06.
Artigo em Inglês | MEDLINE | ID: mdl-31906943

RESUMO

Using an established CRISPR-Cas mediated genome editing technique for streptomycetes, we explored the combinatorial biosynthesis potential of the auroramycin biosynthetic gene cluster in Streptomyces roseosporous. Auroramycin is a potent anti-MRSA polyene macrolactam. In addition, auroramycin has antifungal activities, which is unique among structurally similar polyene macrolactams, such as incednine and silvalactam. In this work, we employed different engineering strategies to target glycosylation and acylation biosynthetic machineries within its recently elucidated biosynthetic pathway. Auroramycin analogs with variations in C-, N- methylation, hydroxylation and extender units incorporation were produced and characterized. By comparing the bioactivity profiles of five of these analogs, we determined that unique disaccharide motif of auroramycin is essential for its antimicrobial bioactivity. We further demonstrated that C-methylation of the 3, 5-epi-lemonose unit, which is unique among structurally similar polyene macrolactams, is key to its antifungal activity.


Assuntos
Antibacterianos/biossíntese , Antifúngicos/química , Vias Biossintéticas/genética , Engenharia Metabólica/métodos , Streptomyces/genética , Antibacterianos/química , Antibacterianos/farmacologia , Antifúngicos/farmacologia , Sistemas CRISPR-Cas , Edição de Genes/métodos , Staphylococcus aureus Resistente à Meticilina/efeitos dos fármacos , Testes de Sensibilidade Microbiana , Polienos/química , Streptomyces/metabolismo
6.
Int J Syst Evol Microbiol ; 70(1): 569-575, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-31647406

RESUMO

Four strains of bacteria designated as AR-3-6T, AT-3-1, AR-3-8T and AR-3-15 were isolated from Arctic soil. Cells were aerobic, Gram-staining-negative, non-motile, non-spore-forming, rod-shaped and yellow-pigmented. Flexirubin-type pigments were present in all strains. All strains tolerated 2 % of NaCl and were psychrotolerant. A phylogenetic analysis based on its 16S rRNA gene sequence revealed that these strains formed a lineage within the family Cytophagaceae that were distinct from various members of the genus Dyadobacter. The closest member of strain AR-3-6T was D. koreensis DSM 19938T (97.2 % sequence similarity) and AR-3-8T was D. hamtensis HHS 11T (97.9 %). The predominant respiratory quinone was menaqinone-7. The major polar lipid was phosphatidylethanolamine. The major cellular fatty acids were summed feature 3 (C16 : 1ω7c and/or C16 : 1ω6c), iso-C15 : 0, C16 : 1ω5c and iso-C17 : 0 3-OH. The DNA G+C content of strains ranges from 40.1 to 42.1 mol%. On the basis of phenotypic, genotypic, chemotaxonomic and phylogenetic analysis, both strains AR-3-6T and AR-3-8T represent a novel member in the genus Dyadobacter, for which the name Dyadobacter psychrotolerans sp. nov. and Dyadobacter frigoris sp. nov. are proposed, respectively. The type strain of Dyadobacter psychrotolerans is AR-3-6T (=KEMB 9005-743T=KACC 21172T=NBRC 113790T) and type strain of Dyadobacter frigoris is AR-3-8T (=KEMB 9005-744T= KACC 21173T=NBRC 113791T).


Assuntos
Cytophagaceae/classificação , Filogenia , Microbiologia do Solo , Regiões Árticas , Técnicas de Tipagem Bacteriana , Composição de Bases , Cytophagaceae/isolamento & purificação , DNA Bacteriano/genética , Ácidos Graxos/química , Fosfatidiletanolaminas/química , Pigmentação , Polienos/química , RNA Ribossômico 16S/genética , Análise de Sequência de DNA , Vitamina K 2/análogos & derivados , Vitamina K 2/química
7.
Chem Commun (Camb) ; 56(5): 822-825, 2020 01 16.
Artigo em Inglês | MEDLINE | ID: mdl-31848534

RESUMO

Produced by a newly isolated Streptomycetes strain, meijiemycin is a gigantic linear polyene-polyol that exhibits structural features not seen in other members of the polyene-polyol family. We propose a biosynthetic mechanism and demonstrate that meijiemycin inhibits hyphal growth by inducing the aggregation of ergosterol and restructuring of the fungal plasma membrane.


Assuntos
Antifúngicos/farmacologia , Álcoois Graxos/farmacologia , Polienos/farmacologia , Antifúngicos/isolamento & purificação , Antifúngicos/metabolismo , Candida albicans/efeitos dos fármacos , Descoberta de Drogas , Álcoois Graxos/isolamento & purificação , Álcoois Graxos/metabolismo , Genes Bacterianos , Genômica , Testes de Sensibilidade Microbiana , Família Multigênica , Polienos/isolamento & purificação , Polienos/metabolismo , Policetídeo Sintases/genética , Streptomyces/química
8.
J Microbiol ; 57(12): 1079-1085, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31758394

RESUMO

A yellow pigmented, Gram-stain-negative, strictly aerobic, rod-shaped, motile by means of gliding, catalase and oxidase positive bacterium, designated strain DS2-AT, was isolated from soil. Growth was observed at 4-32°C (optimum, 28°C), pH 6-9 (optimum, 7.0), and with 0-0.25% (w/v) NaCl (optimum, 0%). Phylogenetic analysis of 16S rRNA gene sequence revealed that strain DS2-AT belonged to the genus Flavobacterium and was most closely related to Flavobacterium aquatile LMG 4008T (96.4%), Flavobacterium terrae DSM 18829T (95.6%), Flavobacterium vireti THG-SM1T (95.5%), Flavobacterium inkyongense IMCC27201T (95.4%), Flavobacterium brevivitae TTM-43T (95.2%), and Flavobacterium cucumis DSM 18830T (95.2%). Strain DS2-AT produces flexirubin-type pigments. The major fatty acids were iso-C15:0, iso-C17:0 3-OH, and iso-C15:0 3-OH. The major respiratory quinone was identified as menaquinone-6. The major polar lipid was found to be phosphatidylethanolamine. The average nucleotide identity values between strain DS2-AT and selected taxa, F. aquatile LMG 4008T, F. terrae DSM 18829T, and F. cucumis DSM 18830T, were 72, 72.7, and 71.6%, respectively. The draft genome of strain DS2-AT has a number of 14 contigs, scaffold N50 of 476,310 bp and a total size of 3,563,867 bp. Additionally, strain DS2-AT contains 3,127 of gene, 41 of tRNA, 6 of rRNA, and 3 of ncRNA. The DNA G + C content of stain DS2-AT was 40.7 mol%. Based on phylogenetic and phenotypic analyses, strain DS2-AT is considered as a novel species of the genus Flavobacterium, for which the name Flavobacterium humi sp. nov., (type strain DS2-AT = KACC 19715T = JCM 32786T) has been proposed.


Assuntos
Flavobacterium/classificação , Flavobacterium/isolamento & purificação , Flavobacterium/metabolismo , Filogenia , Pigmentos Biológicos/metabolismo , Polienos/metabolismo , Microbiologia do Solo , Técnicas de Tipagem Bacteriana , Composição de Bases , DNA Bacteriano/genética , Ácidos Graxos/análise , Flavobacterium/genética , Hibridização de Ácido Nucleico , Fosfatidiletanolaminas/análise , RNA Ribossômico 16S/genética , República da Coreia , Análise de Sequência de DNA , Solo , Vitamina K 2/análogos & derivados , Vitamina K 2/análise
9.
ACS Appl Mater Interfaces ; 11(50): 46591-46603, 2019 Dec 18.
Artigo em Inglês | MEDLINE | ID: mdl-31742377

RESUMO

Oral candidiasis as a highly prevalent and recurrent infection in medically compromised individuals is mainly caused by the opportunistic fungal pathogen Candida albicans. This epithelial infection, if not controlled effectively, can progress to life-threatening systemic conditions and complications. The efficacy of current frontline antifungals is limited due to their poor bioavailability and systemic toxicity. As such, an efficient intervention is essential for controlling disease progression and recurrence. Herein, a theranostic nanoplatform (CD-Gu+-AmB) was developed to track the penetration of antifungals and perturb the invasion of C. albicans at oral epithelial tissues, via decorating the homemade red-emissive carbon dots (CD) with positively charged guanidine groups (Gu+) followed by conjugation with antifungal polyene (amphotericin B, AmB) in a reacting site-controllable manner. The generated CD-Gu+-AmB favorably gathered within the Candida cells and exhibited potent antifungal effects in both planktonic and biofilm forms. It selectively accumulated in the nuclei of human oral keratinocytes and exhibited undetectable toxicity to the host cells. Moreover, we reported for the first time the penetration and exfoliation profiles of CD in a three-dimensional organotypic model of human oral epithelial tissues, demonstrating that the extra- and intracellular accumulation of CD-Gu+-AmB effectively resisted the invasion of C. albicans by forming a "shielding" layer throughout the entire tissue. This study establishes a multifunctional CD-based theranostic nanoplatform functioning as a traceable and topically applied antifungal to arm oral epithelia, thereby shedding light on early intervention of mucosal candidiasis for oral and general health.


Assuntos
Anfotericina B/farmacologia , Antifúngicos/farmacologia , Candidíase/tratamento farmacológico , Infecções Fúngicas Invasivas/tratamento farmacológico , Anfotericina B/química , Antifúngicos/química , Biofilmes/efeitos dos fármacos , Disponibilidade Biológica , Candida albicans/efeitos dos fármacos , Candida albicans/patogenicidade , Candidíase/microbiologia , Carbono/química , Células Epiteliais/efeitos dos fármacos , Células Epiteliais/microbiologia , Guanosina Monofosfato/química , Humanos , Infecções Fúngicas Invasivas/microbiologia , Queratinócitos/efeitos dos fármacos , Testes de Sensibilidade Microbiana , Mucosa Bucal/efeitos dos fármacos , Mucosa Bucal/microbiologia , Polienos/química , Polienos/farmacologia , Pontos Quânticos/química
10.
J Mass Spectrom ; 54(10): 823-833, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31476245

RESUMO

Actinobacteria are one of the most promising producers of medically and industrially relevant secondary metabolites. However, screening of such compounds in actinobacteria growth demands simple, fast, and efficient extraction procedures that enable detection and precise quantification of biologically active compounds. In this regard, solid phase microextraction (SPME) emerges as an ideal extraction technique for screening of secondary metabolites in bacteria culture due to its non-exhaustive, minimally invasive, and non-destructive nature: its integrated sample preparation workflow; balanced coverage feature; metabolism quenching capabilities; and superior cleanup, as well as its versatility in configuration, which enables automation and high throughput applications. The current work provides a comparison of micro-scale and direct immersion SPME (DI-SPME) for screening of secondary metabolites, describes the optimization of the developed DI-SPME method, and introduces the developed technique for mapping of target secondary metabolites as well as its direct coupling to mass spectrometry for such applications. The optimized DI-SPME method provided higher amounts of extracted ions and intensity signals, yielding superior extraction and desorption efficiency as compared with micro-scale extraction. Studied compounds presented stability on the coating for 24 h at room temperature. The DI-SPME mapping approach revealed that lysolipin I and the lienomycin analog are distributed along the center and edges of the colony, respectively. Direct coupling of SPME to MS provided a similar ions profile as SPME-LC-MS while enabling a significant decrease in analysis time, demonstrating its suitability for such applications. DI-SPME is herein presented as an alternative to micro-scale extraction for screening of secondary metabolites in actinobacteria solid medium, as well as a feasible alternative to DESI-IMS for mapping of biologic radial distribution of secondary metabolites and cell life cycle studies. Lastly, the direct coupling of DI-SPME to MS is presented as a fast, powerful technique for high throughput analysis of secondary metabolites in this medium.


Assuntos
Actinobacteria/metabolismo , Metabolômica , Metabolismo Secundário , Microextração em Fase Sólida , Cromatografia Líquida de Alta Pressão , Ensaios de Triagem em Larga Escala , Polienos/análise , Análise de Componente Principal , Espectrometria de Massas em Tandem , Xantenos/análise
11.
Nat Chem ; 11(10): 906-912, 2019 10.
Artigo em Inglês | MEDLINE | ID: mdl-31548673

RESUMO

Polyketide synthases assemble diverse natural products with numerous important applications. The thioester intermediates in polyketide assembly are covalently tethered to acyl carrier protein domains of the synthase. Several mechanisms for polyketide chain release are known, contributing to natural product structural diversification. Here, we report a dual transacylation mechanism for chain release from the enacyloxin polyketide synthase, which assembles an antibiotic with promising activity against Acinetobacter baumannii. A non-elongating ketosynthase domain transfers the polyketide chain from the final acyl carrier protein domain of the synthase to a separate carrier protein, and a non-ribosomal peptide synthetase condensation domain condenses it with (1S,3R,4S)-3,4-dihydroxycyclohexane carboxylic acid. Molecular dissection of this process reveals that non-elongating ketosynthase domain-mediated transacylation circumvents the inability of the condensation domain to recognize the acyl carrier protein domain. Several 3,4-dihydroxycyclohexane carboxylic acid analogues can be employed for chain release, suggesting a promising strategy for producing enacyloxin analogues.


Assuntos
Antibacterianos/biossíntese , Polienos/metabolismo , Policetídeo Sintases/metabolismo , Acinetobacter baumannii/efeitos dos fármacos , Acilação , Antibacterianos/química , Antibacterianos/farmacologia , Testes de Sensibilidade Microbiana , Estrutura Molecular , Polienos/química , Polienos/farmacologia
12.
Nat Chem ; 11(10): 913-923, 2019 10.
Artigo em Inglês | MEDLINE | ID: mdl-31548674

RESUMO

Modular polyketide synthases and non-ribosomal peptide synthetases are molecular assembly lines that consist of several multienzyme subunits that undergo dynamic self-assembly to form a functional megacomplex. N- and C-terminal docking domains are usually responsible for mediating the interactions between subunits. Here we show that communication between two non-ribosomal peptide synthetase subunits responsible for chain release from the enacyloxin polyketide synthase, which assembles an antibiotic with promising activity against Acinetobacter baumannii, is mediated by an intrinsically disordered short linear motif and a ß-hairpin docking domain. The structures, interactions and dynamics of these subunits were characterized using several complementary biophysical techniques to provide extensive insights into binding and catalysis. Bioinformatics analyses reveal that short linear motif/ß-hairpin docking domain pairs mediate subunit interactions in numerous non-ribosomal peptide and hybrid polyketide-non-ribosomal peptide synthetases, including those responsible for assembling several important drugs. Short linear motifs and ß-hairpin docking domains from heterologous systems are shown to interact productively, highlighting the potential of such interfaces as tools for biosynthetic engineering.


Assuntos
Peptídeo Sintases/química , Polienos/química , Policetídeo Sintases/química , Cristalografia por Raios X , Simulação de Acoplamento Molecular , Peptídeo Sintases/metabolismo , Polienos/metabolismo , Policetídeo Sintases/metabolismo , Conformação Proteica
13.
Waste Manag ; 100: 18-27, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31494403

RESUMO

The decrease in stress crack resistance (SCR) of geomembranes (GMBs) due to physical ageing is examined. It is shown that the SCR of a 1.5 mm high density polyethylene (HDPE) GMB immersed in municipal solid waste (MSW) leachate, deionized (DI) water, and air experienced a decrease to an equilibrium value, denoted herein as SCRm, in a few months and then did not change for the remainder of a 116-month test. The same GMB was also immersed in basic mining pregnant leach solution (PLS) and brine and a similar decrease in SCR to SCRm was observed in the first three months. For this GMB, SCRm was about 50% of the initial SCR value (i.e., SCRm ∼ 0.5·SCRo) for all incubation fluids examined. It is then shown that this behaviour is not unique. Seven other high density polyethylene (HDPE) and one blended polyolefin (BPO) GMBs are shown to experience a decrease in SCR to 0.17·SCRo ≤ SCRm ≤ 0.56·SCRo without any evidence of oxidative degradation. The magnitude of SCRm is shown to be product specific. In contrast, the one linear low density polyethylene (LLDPE) examined exhibited no such physical ageing. The implications for designing with GMBs are discussed.


Assuntos
Eliminação de Resíduos , Gerenciamento de Resíduos , Polienos , Polietileno , Resíduos Sólidos , Instalações de Eliminação de Resíduos
14.
Waste Manag ; 98: 160-172, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31450178

RESUMO

Sorting at industrial scale is required to perform mechanical recycling of plastics in order to obtain properties that could be competitive with virgin polymers. As a matter of fact, the most part of the various types of plastic waste are not miscible and even compatible. Mid-Infrared (MIR) HyperSpectral Imagery (HSI) is viewed as one of the solutions to the problem of black plastic sorting. Many Waste of Electrical and Electronic Equipment (WEEE) plastics are black. Nowadays, these materials are difficult to sort at an industrial scale because the main used pigment to produce this color, carbon black, masks the Near-Infrared (NIR) spectra of polymers, the currently most used technology for acute sorting in industrial conditions. In this study, laboratory Fourier-Transform Infrared (FTIR) in Attenuated Total Reflection mode (ATR) has been used as a theoretical toolbox based on physical chemistry to help building an automated HSI discrimination despite its limited conditions, especially shorter wavelengths ranges. Weaker resolution and very short acquisition times are other HSI limitations. Helping fast and exhaustive laboratory characterizations of polymeric waste stocks is the other goal of this study. This study focusses on polyolefins as they represent the second biggest fraction of WEEE plastics (WEEP) after styrenics and since little quantities mixed to styrenics during mechanical recycling can lead to important decrease in mechanical properties. Twelve references were thus evaluated and compared between each other and with real waste samples to highlight spectral elements, which can enable differentiation. Charts compiling the signals of discussed polymers were built aiming to the same objective.


Assuntos
Resíduo Eletrônico , Plásticos , Polienos , Polímeros , Reciclagem
15.
Plant Mol Biol ; 101(3): 297-313, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31368003

RESUMO

KEY MESSAGE: An enzyme is crucial for the formation of Hedychium coronarium scent and defense responses, which may be responsible for the biosynthesis of allo-ocimene in H. coronarium. Hedychium coronarium can emit a strong scent as its main scent constituents are monoterpenes and their derivatives. Among these derivatives, allo-ocimene is not only a very important volatile substance in flower aroma, but is also crucial to plant defense. However, the molecular mechanism of allo-ocimene biosynthesis has not been characterized in plants. In this study, a new alcohol dehydrogenase gene, HcADH, was cloned. The amino acid sequences encoded by HcADH contained the most conserved motifs of short chain alcohol dehydrogenase/reductases (SDRs), which included NAD+ binding domain, TGxxx[AG]xG and active site YxxxK. Real-time PCR analyses showed that the HcADH was highly expressed in the outer labellum but was almost undetectable in vegetative organs. The change in its expression level in petals was positively correlated with the emission pattern of allo-ocimene during flower development. HcADH expression coincides also the release level of allo-ocimene among different Hedychium species. Although HcADH is not expressed in the leaves, HcADH expression and allo-ocimene release in leaves can be induced by mechanical wounding or methyl jasmonate (MeJA) treatment. In addition, the expression of HcADH induced by mechanical wounding can be prevented by acetylsalicylic acid, a jasmonic acid biosynthesis inhibitor, suggesting that jasmonic acid might participate in the transmission of wounding signals. Using the Barley stripe mosaic virus (BSMV)-VIGS method, it was found that BSMV:HcADH335 inoculation was able to down-regulate HcADH expression, decreasing only the release of allo-ocimene in flowers while the content of other volatile substances did not decrese. In vitro characterization showed that recombinant HcADH can catalyze geraniol into citral, and citral is an intermediate of allo-ocimene biosynthesis. HcADH may be responsible for the biosynthesis of allo-ocimene in H. coronarium, which is crucial for the formation of H. coronarium scent and defense function.


Assuntos
Proteínas de Plantas/metabolismo , Polienos/metabolismo , Redutases-Desidrogenases de Cadeia Curta/metabolismo , Zingiberaceae/enzimologia , Acetatos/metabolismo , Monoterpenos Acíclicos , Ciclopentanos/metabolismo , Flores/enzimologia , Perfilação da Expressão Gênica , Regulação da Expressão Gênica de Plantas , Oxilipinas/metabolismo , Proteínas de Plantas/genética , Redutases-Desidrogenases de Cadeia Curta/genética , Transdução de Sinais , Terpenos/metabolismo , Zingiberaceae/genética
16.
J Med Microbiol ; 68(9): 1353-1358, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31271350

RESUMO

Introduction. Candida auris is a pathogenic yeast that mainly affects immunosuppressed patients and those with implanted medical devices. This pathogen also displays elevated resistance to common antifungals and high survival and spreading capacities. Since no antifungal breakpoints have yet been defined for this pathogen, the data obtained here can be useful for further research concerning treatment or implementation of a prevention and disinfection protocol. Our aim was to study the antifungal resistance of C. auris to current antifungals in planktonic and sessile states. Using confocal laser scanning microscopy and viable biomass production, we demonstrated the ability of C. auris to develop a mature biofilm. We compared the minimal inhibitory concentration (MIC) and the minimal biofilm eradication concentration (MBEC) for the C. auris DSM 21092 strain plus two clinical isolates, and the results were compared with those obtained for Candida albicans and Candida parapsilosis, two species strongly linked to bloodstream infections and infections associated with biomaterials. We found that the clinical isolates of C. auris were resistant to fluconazole and sensitive to echinocandins and polyenes. The C. auris biofilms did not show susceptibility to any antifungal agent, showing MBECs that were up to 512-fold higher than the MICs. These findings highlight the importance of biofilm formation as a key factor underlying the resistance of this species to antifungals and suggest that the presence of implantable medical devices is one of the major risk factors in immunocompromised patients.


Assuntos
Antifúngicos/farmacologia , Biofilmes/efeitos dos fármacos , Candida/efeitos dos fármacos , Candida/isolamento & purificação , Candida albicans/efeitos dos fármacos , Candida parapsilosis/efeitos dos fármacos , Candidíase/microbiologia , Contagem de Colônia Microbiana , Farmacorresistência Fúngica , Equinocandinas/farmacologia , Fluconazol/farmacologia , Humanos , Testes de Sensibilidade Microbiana , Microscopia Confocal , Polienos/farmacologia
17.
Molecules ; 24(13)2019 Jul 02.
Artigo em Inglês | MEDLINE | ID: mdl-31269674

RESUMO

Composites of polyolefin matrices (HDPE and PP) were prepared by melt processing using two commercially available nano ZnO powders (Zinkoxyd aktiv and Zano 20). The mechanical and thermal properties, UV-Vis stability, and antibacterial activity of composites were studied. Tensile testing revealed that both nano ZnO types have no particular effect on the mechanical properties of HDPE composites, while some positive trends are observed for the PP-based composites, but only when Zano 20 was used as a nanofiller. Minimal changes in mechanical properties of composites are supported by an almost unaffected degree of crystallinity of polymer matrix. All polyolefin/ZnO composites exposed to artificial sunlight for 8-10 weeks show more pronounced color change than pure matrices. This effect is more evident for the HDPE than for the PP based composites. Color change also depends on the ZnO concentration and type; composites with Zano 20 show more intense color changes than those prepared with Zinkoxyd aktiv. Results of the antibacterial properties study show very high activity of polyolefin/ZnO composites against Staphylococcus aureus regardless of the ZnO surface modification, while antibacterial activity against Escherichia coli shows only the composites prepared with unmodified ZnO. This phenomenon is explained by different membrane structure of gram-positive (S. aureus) and gram-negative (E. coli) bacteria.


Assuntos
Química Orgânica/métodos , Polienos/química , Óxido de Zinco/química , Antibacterianos/farmacologia , Escherichia coli/efeitos dos fármacos , Testes de Sensibilidade Microbiana , Nanopartículas/química , Nanopartículas/ultraestrutura , Silanos/química , Espectroscopia de Infravermelho com Transformada de Fourier , Staphylococcus aureus/efeitos dos fármacos , Ácidos Esteáricos/química , Propriedades de Superfície , Resistência à Tração , Difração de Raios X
18.
J Infect Chemother ; 25(10): 743-749, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31257156

RESUMO

Candida auris is a multidrug-resistant and emergent pathogen that has caused healthcare-associated infection outbreaks. Recently, C. auris has spread worldwide; nevertheless, it was unexpectedly rare before 2009. Based on the molecular epidemiological analysis, C. auris may independently emerge at specific areas at first and recently may be transmitted to other continents. As C. auris cannot be detected using conventional methods, internally transcribed spacers, D1/D2 regions of the 26S rDNA sequencing, and/or matrix-assisted laser desorption ionization time-of-flight mass spectrometry method can be selected as comparatively accessible choices. Thus, detection of C. auris using the conventional method might be underestimated. In Japan, all C. auris strains were isolated from ear specimen and not from invasive mycoses. Japan strains were classified as an East Asian clade under a single clone. Although colonization, virulence, and infection pattern are almost the same as with other Candida species, its antifungal resistance is different. Fluconazole resistance is notably common, but resistance to all three classes of antifungals (azole, polyene, and echinocandin) rarely exists. Once C. auris is detected, screening, emphasis on hand hygiene adherence, use of single-patient room isolation, contact precaution, surveillance, and eradication from the environment and patients are appropriately required for infection control.


Assuntos
Antifúngicos/farmacologia , Candida/isolamento & purificação , Candidíase/tratamento farmacológico , Antifúngicos/uso terapêutico , Azóis/farmacologia , Azóis/uso terapêutico , Candida/efeitos dos fármacos , Candida/patogenicidade , Candidíase/epidemiologia , Candidíase/microbiologia , Farmacorresistência Fúngica Múltipla , Equinocandinas/farmacologia , Equinocandinas/uso terapêutico , Fluconazol/farmacologia , Fluconazol/uso terapêutico , Humanos , Japão/epidemiologia , Testes de Sensibilidade Microbiana , Polienos/farmacologia , Polienos/uso terapêutico , Prevalência
19.
Carbohydr Polym ; 222: 115014, 2019 Oct 15.
Artigo em Inglês | MEDLINE | ID: mdl-31320045

RESUMO

The films based on carboxymethyl derivatives of starch (CMS) and cellulose (CMC) were proposed as a novel human skin equivalent. The physicochemical properties (moisture absorption, solubility in water, mechanical properties) of CMS/CMC films were evaluated. Additionally, some properties were compared to the human skin ones (surface roughness, tribology). The system based on CMS/CMC 25/75 wt. % was selected for testing the adhesive properties of pressure-sensitive adhesives commonly used for medical purposes (acrylic, silicone and polyisobutylene). Similar tests were performed for human skin. The peel adhesion values for CMS/CMC film and human skin were similar for all adhesives types tested. Applying such a skin equivalent allows to evaluate the functional properties of medical pressure-sensitive adhesives without the ethical and economic issues.


Assuntos
Adesivos/química , Carboximetilcelulose Sódica/química , Pele Artificial , Amido/análogos & derivados , Resinas Acrílicas/química , Adesividade , Humanos , Interações Hidrofóbicas e Hidrofílicas , Polienos/química , Polímeros/química , Silicones/química , Amido/química , Resistência à Tração
20.
Macromol Rapid Commun ; 40(19): e1900302, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31334910

RESUMO

Adamantyl vinyl ketone (AVK) and its copolymers are synthesized using reversible addition fragmentation chain-transfer (RAFT) methodology and then degraded using UV light. The polymerization of AVK is found to be controlled as indicated by a linear correlation between the molecular weights of the polymers produced and monomer conversion as well as a series of chain extensions. The RAFT method is also used to synthesize random and block copolymers of AVK and methyl methacrylate. Irradiating poly(adamantyl vinyl ketone) (PAVK) with UV light affords a polyolefin and adamantane as the major products. Similar products are obtained, along with poly(methyl methacrylate) (PMMA), when the block copolymer is subjected to UV light. The random copolymer undergoes complete degradation under similar conditions. A mechanism wherein stereoelectronic effects channel photodegradation through Norrish I Type pathways in a manner that preserves the main chain of the polymer during the decomposition process is proposed.


Assuntos
Adamantano/síntese química , Polienos/síntese química , Adamantano/análogos & derivados , Adamantano/química , Estrutura Molecular , Processos Fotoquímicos , Polienos/química , Estereoisomerismo , Raios Ultravioleta
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