The present work describes a chemical study, monitored by the antimicrobial activity, from the following plant species from Alagoas flora: Zeyheria tuberculosa (Vell.) Bur. (Bignoniaceae), Capparis cynophallophora L. and C. spinosa Jacq (Capparaceae), Swatzia apetala Raddi. (Fabaceae), Piper marginatum Jacq. and P. hayneanum C.DC. (Piperaceae), and Portulaca cf. elation Mart. ex. Rohrb. (Portulacaceae). The antimicrobial evaluation of the extracts, fractions and subfractions was carried out by the Agar diffusion method, and the bioprospection with isolated substances was investigated by the bioautographic assay. The plant species were tested against a panel of microorganisms, including one Gram positive bacteria Staphylococcus aureus (ATCC 25923), one Gram negative Pseudomonas aeruginosa (ATCC 27853), and one fungus Candida albicans (ATCC 10231). The Minimum Inhibitory Concentration (MIC) was determined for the fractions and subfractions that showed inhibitory activity against the tested microorganisms. Only the material extracted from the species Z. tuberculosa and P. hayneanum were able to inhibt S. aureus and C. albicans growth. These microorganisms were sensible to CHCl3 subfractions from Z. tuberculosa stem and P. hayneanum stem and roots, with MIC from 1 to 250 μg/disc. Active subfractions, submitted to a bioassay with the brine shrimp Artemia salina, presented low or no toxicity at all. Pomade made out of subfracionated extracts CHCl3-MeOH (1:1) and CHCl3-AcOEt (1:1) from P. hayneanum showed a good potential for antimicrobial application in in vivo assays for infected wound treatment in rats. The phytochemical investigation from Z. tuberculosa extracts with antimicrobial potential, led to the isolation of four flavonoids (5,6,7,8-Tetramethoxyflavone 5,6,7-Trimethoxyflavone, 4’-Hydroxy-5,6,7,8-Tetramethoxyflavone and 4’ -Hidroxi-5,6,7-tetramethoxyflavone); while in the P. hayneanum species three phytosteroids were isolated (β-Sitosterol, Stigmasterol and β-Sitosterol-Dglucopiranoside). These substances were identified from analysis of RMN spectroscopics uni and bidimensional (HMBC e HSQC), including DEPT. The phytochemical study contributed to extend the chemical taxonomic profile from Zeyheria gender. The isolated substances tested by bioautography method with antimicrobial activity against S. aureus were 5,6,7,8-Tetramethoxyflavone, 5,6,7-Trimethoxyflavone and 4’-Hydroxy-5,6,7,8-tetramethoxyflavone, while against C. albicans were 5,6,7,8- Tetramethoxyflavone, 4’-Hydroxy-5,6,7,8-tetramethoxyflavone and β-Sitosterol-3-O-β-D-glucopiranoside. The results are considered promising, since that the extracts, fractions and isolated compounds showed ability to control the growth of microorganisms responsible for contamination of wounds that affect the integumentary system. / Fundação de Amparo a Pesquisa do Estado de Alagoas / Conselho Nacional de Desenvolvimento Científico e Tecnológico / O presente trabalho descreve o estudo químico, monitorado pela atividade antimicrobiana, das espécies vegetais da flora alagoana: Zeyheria tuberculosa (Vell.) Bur. (Bignoniaceae), Capparis cynophallophora L e C. spinosa Jacq (Capparaceae), Swatzia apetala Raddi. (Papilionaceae), Piper marginatum Jacq. e P. hayneanum C.DC. (Piperaceae), e Portulaca cf. elation Mart. ex. Rohrb. (Portulacaceae). A avaliação da potencial antimicrobiano dos extratos, frações e subfrações foi realizado pelo método de difusão em ágar, e a bioprospecção com as substâncias isoladas investigado pelo ensaio da bioautografia. As espécies vegetais foram testadas frente a um painel de microrganismos, incluindo uma bactéria Gram-positiva Staphylococcus aureus (ATCC 25923), uma Gram-negativa Pseudomonas aeruginosa (ATCC 27853), e um fungo Candida albicans (ATCC 10231). A Concentração Inibitória Mínima (CIM) foi determinada para as frações e subfrações que apresentaram atividade inibitória sobre os microrganismos testados. Somente material extraído das espécies vegetais Z. tuberculosa e P. hayneanum foram capazes de inibir o crescimento do S. aureus e da C. albicans. Estes microrganismos foram mais sensíveis à ação das subfrações em CHCl3 oriundas do caule de Z. tuberculosa, e do caule e das raízes de P. hayneanum, com CIM de 1 a 250 μg/disco. As subfrações ativas, submetidas ao bioensaio com o microcrustáceo Artemia salina, apresentaram baixa toxicidade ou foram atóxicas. Pomadas a base de extratos subfracionados CHCl3-MeOH (1:1) e CHCl3-AcOEt (1:1) de P. hayneanum mostraram bom potencial de aplicação como antimicrobiano em ensaios in vivo para tratamento de feridas infectadas em ratos. A investigação fitoquímica de extratos de Z. tuberculosa com potencial antimicrobiano conduziu ao isolamento de quatro flavonóides (5,6,7,8-Tetrametoxiflavona, 5,6,7-Trimetoxiflavona, 4’ -Hidroxi-5,6,7,8-tetrametoxiflavona e 4’ -Hidroxi-5,6,7-trimetoxiflavona); enquanto que da espécie P. hayneanum foram isolados três fitoesteróides (β-Sitosterol, Estigmasterol e β-Sitosterol glicosilado). Estas substâncias foram identificadas com base na análise de dados espectroscópicos de RMN uni e bidimensional (HMBC e HSQC), incluindo DEPT. O estudo fitoquímico contribuiu para ampliar o perfil quimiotaxonômico do gênero Zeyheria. Nos ensaios antimicrobianos com as substâncias isoladas pelo método da bioautografia as substâncias ativas contra S. aureus foram 5,6,7,8-Tetrametoxiflavona, 5,6,7-Trimetoxiflavona e 4’ -Hidroxi-5,6,7,8-tetrametoxiflavona, enquanto para C. albicans foram 5,6,7,8-Tetrametoxiflavona, 4’ -Hidroxi-5,6,7,8-tetrametoxiflavona e 3-O-β-D-Glicopiranosilsitosterol. Os resultados obtidos são considerados promissores, uma vez que, os extratos, frações e compostos isolados apresentaram capacidade de controlar o crescimento de microrganismos responsáveis pela contaminação de feridas que afetam o sistema tegumentar.

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior / The modern dentistry emphasizes the importance of the control of the dental biofilm to improve the oral health. The use in the odontologic therapeutics rinses has a crucial paper in the reduction of the biofilm. The effectiveness of these substances was appraised extensively in several studies. The pharmacologic research of medicinal plants is necessary due to its therapeutic capacity and use. Essential oils and extracts of plants have been used for several applications in the popular medicine, among them as antimicrobial and antiseptic topics. There are several methods described inside the literature for size the antimicrobial activity of the essential oils. However the measure of this activity is hindered by the absence of standardized methods, what dificults the comparison among the studies. Besides, disables its reproduction. The present work researched in vitro the susceptibility of Streptococcus mutans to the Hyptis pectinata essential oil. It presents antimicrobial activity equivalent to the chlorhexidine, what demonstrates the use of this oil as promising alternative in the regiments of oral health. Validating the antimicrobial properties of the Hyptis pectinata essential oil, an experiment was developed using the diffusion technique in disk with some variations in the diluents of the oil, among them Tween 20 and Tween 80 are the most suitable for the emulsification of the Hyptis pectinata oil. The obtained data emphasize a considerable interest for the development of reliable methods for evaluation and comparison of the activity antimicrobial of essential oils. / A odontologia moderna enfatiza a importância do controle do biofilme dental para melhorar a saúde oral. O uso na terapêutica odontológica de colutórios tem um papel crucial na redução do biofilme. A eficácia destas substâncias foi extensivamente avaliada em vários estudos. A pesquisa farmacológica de plantas medicinais é necessária diante de sua capacidade terapêutica e da sua larga utilização. Óleos essenciais e extratos de plantas há muito tempo têm sido base para diversas aplicações na medicina popular, entre elas como antimicrobianos e anti-sépticos tópicos. Existem vários métodos descritos na literatura para mensurar a atividade antimicrobiana dos óleos essenciais. Entretanto, a ausência de métodos padronizados, faz com que a comparação entre os estudos seja praticamente impossível, além de incapacitar sua reprodutibilidade. O presente trabalho pesquisou in vitro a eficácia do óleo essencial Hyptis pectinata frente ao Streptococcus mutans. Este apresenta atividade antimicrobiana equivalente à clorexidina, o que demonstra o uso deste óleo como alternativa promissora nos regimentos de saúde oral. Validando as propriedades antimicrobianas do óleo essencial de Hyptis pectinata, desenvolveu-se um experimento utilizando a técnica de difusão em disco, com algumas variações nos diluentes do óleo, dos quais, o Tween 20 e o Tween 80 são os mais indicados para a emulsificação do óleo de Hyptis pectinata. Os dados obtidos ressaltam um considerável interesse pelo desenvolvimento de métodos confiáveis para avaliação e comparação da atividade antimicrobiana de óleos essenciais.

Orientadores: Fabiana Fantinatti Garboggini, Marta Cristina Teixeira Duarte / Dissertação (mestrado) - Universidade Estadual de Campinas, Instituto de Biologia / Made available in DSpace on 2018-08-26T21:03:15Z (GMT). No. of bitstreams: 1 Franca_Paula_M.pdf: 7905078 bytes, checksum: 6735b6d77cec7c206090abaa6e56fdfc (MD5) Previous issue date: 2015 / Resumo: Os micro-organismos associados ao continente Antártico apresentam populações diversas e metabolicamente ativas, porém o potencial farmacológico dos compostos obtidos pelos micro-organismos é pouco conhecido. As bactérias são importante fonte de compostos utilizados atualmente como antimicrobianos, e as principais vias de biossíntese de muitos antibióticos são catalisadas pelos genes PKS (Polyketide Synthases) e NRPS (Non Ribosomal Peptide Synthetases). O presente estudo teve como objetivo a avaliação genética e funcional da produção de compostos antimicrobianos obtidos de bactérias isoladas na Baía do Almirantado, Antártica, a identificação taxonômica das bactérias que apresentaram tal potencial e a identificação do perfil químico dos compostos antimicrobianos. O total de 153 bactérias foi isolado do ambiente antártico, 127 isolados apresentaram pelo menos um dos genes PKSI, PKSII e NRPS. Estes foram identificados através do sequenciamento parcial do gene RNA ribossomal 16S e pertencem a 28 gêneros distintos, sendo representantes dos Filos Proteobacteria, Firmicutes, Actinobacteria e Bacteroidetes. A avaliação funcional da produção de antimicrobianos foi realizada com o total de 76 isolados dos quais 30 isolados formaram halos de inibição frente a micro-organismos testes. Os extratos brutos obtidos foram avaliados quanto à concentração inibitória mínima (MIC) frente a oito micro-organismos não virulentos, e 18 extratos brutos apresentaram atividade inibitória, onde se destaca o extrato denominado E131, obtido da bactéria do gênero Streptomyces, que apresentou atividade bacteriostática frente S. aureus e M. luteus, e atividade bactericida frente a C. albicans e B. subtilis. Frente a micro-organismos isolados de amostras clínicas, 11 extratos brutos apresentaram atividade inibitória frente a quatro cepas de Neisseria meningitides, com MIC dos extratos brutos variando de 0,0313 mg.mL-1 até 2,0 mg.mL-1. O extrato E46, obtido da bactéria Pseudoalteromonas sp., destacou-se quanto à atividade inibitória observada frente às quatro cepas avaliadas. Frente à cepa B4, destacam-se a atividade antimicrobiana dos extratos brutos obtido das bactérias Pseudoalteromonas sp. e Pseudomonas azotoformans CUG12536. Frente à cepa YUSA, destaca-se a atividade antimicrobiana observada pelo extrato bruto obtido da bactéria Marinilactibacillus sp., cuja atividade antimicrobiana foi relatada pela primeira vez neste gênero de bactéria. Os extratos brutos possuem em sua composição, compostos cuja atividade antimicrobiana é conhecida, como ácidos graxos e compostos cuja atividade antimicrobiana não está descrita na literatura. Os testes de fracionamento dos extratos frente a solventes de diferentes polaridades indicou que os extratos brutos são solúveis, em sua maioria, a solvente polar. Portanto, as bactérias isoladas da Antártica produzem compostos de interesse farmacológico e podem ser utilizadas como fonte de novos compostos. Tais resultados enfatizam a necessidade de mais estudos de bactérias associadas a ambientes extremos, como a Antártica / Abstract: Microorganisms associated with the Antarctic continent have various and metabolically active populations, but the pharmacological potential of compounds obtained by micro-organisms is poorly understood. Bacterias are an important source of compounds currently used as antimicrobial, major biosynthetic pathways of many antibiotics are catalyzed by the PKS genes (Polyketide Synthases) and NRPS (Non Ribosomal Peptide Synthetases). This study had the objective to genetic and functional evaluation of the antimicrobial activity of bacteria isolated in Admiralty Bay, Antarctica, and the taxonomic identification of bacteria that had such potential and identification of the chemical profile of antimicrobial compounds. The total of 153 bacteria was isolated from Antarctic environment, among the isolates, 127 isolates showed at least one of the genes PKSI, PKSII and NRPS. These were identified by partial sequencing of 16S ribosomal RNA gene and belong to 28 different genera, with representatives of the phyla Proteobacteria, Firmicutes, Actinobacteria and Bacteroidetes. The functional evaluation of the production of antibiotic was conducted with a total of 76 isolates including 30 isolates that formed inhibition halos against test microorganisms. The crude extracts were evaluated as the minimum inhibitory concentration (MIC) against eight non-virulent microorganisms, and 18 crude extracts showed activity, highlighting the so-called E131 extract, obtained from bacteria of the genus Streptomyces, which showed bacteriostatic activity against S. aureus and M. luteus, and bactericidal activity against C. albicans and B. subtilis. Faced with microorganisms isolated from clinical samples, 11 crude extracts showed inhibitory activity against four strains of Neisseria meningitides, with MIC ranging from 0.0313 mg.mL-1 to 2.0 mg.mL-1. The E46 extract obtained from the bacterium Pseudoalteromonas sp., stood out as the inhibitory activity observed across the four evaluated strains. Faced with the strain B4, stand out the antimicrobial activity of crude extracts obtained from bacteria Pseudoalteromonas sp. and Pseudomonas azotoformans CUG12536. Against YUSA strain, Marinilactibacillus sp. crude extract showed antimicrobial activity, which was the first reported in this bacterial genus. The extracts have in their composition, antimicrobial compounds whose activity is known, such as fatty acids, and compounds whose antimicrobial activity is not described in the literature. Fractionation tests of extracts using solvents of different polarities, indicated that the crude extracts are soluble mostly the polar solvent. Therefore, the bacteria isolated from the Antarctic produce compounds of pharmacological interest and can be used as a source of novel compounds. These results highlight the need for more studies of bacteria associated with extreme environments, such as Antarctica / Mestrado / Microbiologia / Mestra em Genética e Biologia Molecular

Orientadores: Marta Cristina Teixeira Duarte, Guiomar Denisse Posada Izquierdo, Fernando Pérez Rodríguez / Dissertação (mestrado) - Universidade Estadual de Campinas, Faculdade de Engenharia de Alimentos / Made available in DSpace on 2018-08-27T10:14:20Z (GMT). No. of bitstreams: 1 Possas_AriciaMaraMelo_M.pdf: 2038081 bytes, checksum: 8baf91fa5300fdd80940e18d382b2468 (MD5) Previous issue date: 2015 / Resumo: Salmonella é um dos patógenos mais importantes relacionados a surtos de origem alimentar que têm como veículos carnes vendidas prontas para o consumo. A contaminação destes produtos pode ocorrer durante o processamento em operações como o fatiamento, embalagem e outros procedimentos que requerem manipulação. O aumento do consumo de produtos à base de carne de peru tem sido verificado mundialmente, pois estes aliam a praticidade dos chamados produtos "ready-to-eat" com o fato de que as carnes de aves são reconhecidas como saudáveis. Outro fator importante que influencia na escolha do consumidor é a presença de conservantes químicos nos produtos alimentícios. A substituição de conservantes químicos por substâncias naturais, como os óleos essenciais, e a incorporação destas em sistemas de embalagens ativas, representa uma alternativa interessante e atende a demanda por produtos mais naturais. Este estudo teve como objetivo desenvolver e aplicar um revestimento incorporado com óleo essencial como antimicrobiano em produtos cárneos à base de peito de peru e avaliar sua atividade contra Salmonella Enteritidis, simulando um sistema de embalagem ativa. Além disso, foi estudada a transferência desta bactéria a produtos de peito de peru durante o fatiamento, para simular a contaminação cruzada em equipamentos utilizados para este procedimento. Também foi avaliado o comportamento de S. Enteritidis durante a vida útil de fatias contaminadas dos produtos cárneos, contendo ou não revestimento de óleo essencial, armazenadas a diferentes temperaturas (10 e 25 °C). Quatro formulações dos produtos cárneos (A, B, C e D) foram estudadas para que fosse possível avaliar como a composição da matriz alimentar influencia no comportamento de S. Enteritidis. Dos quatro óleos essenciais testados in vitro, o óleo essencial de Thymus vulgaris (tomilho) foi o que apresentou menor concentração mínima inibitória (MIC) contra S. Enteritidis, sendo selecionado para ensaios posteriores. Baseando-se em análise sensorial e nos testes de triagem antimicrobiana, uma concentração de 0,8 % (v/m) de óleo essencial de T. vulgaris foi aplicada nos revestimentos para os produtos cárneos. Observou-se transferência de S. Enteritidis durante o fatiamento, a partir das lâminas de corte artificialmente contaminadas (~ 8 log UFC/lâmina) a todas as fatias analisadas (n = 20), de produtos revestidos ou não com emulsão de óleo essencial de T. vulgaris. Diferenças estatísticas foram detectadas em relação à transferência do patógeno nos diferentes tratamentos (p ? 0,05). O comportamento de S. Enteritidis nas fatias durante o armazenamento foi fortemente dependente da temperatura e da composição dos produtos, sendo verificado o crescimento a 25 °C em todas as formulações e tratamentos avaliados. Apenas o produto A propiciou o crescimento do patógeno a 10 °C. Nas fatias dos demais produtos foi verificado um perfil de sobrevivência, com redução das contagens ao longo da vida útil. O óleo essencial de T. vulgaris foi efetivo contra S. Enteritidis, durante o armazenamento de fatias revestidas a 10 °C. Testes finais indicaram que a adição e mistura do óleo essencial juntamente com os outros ingredientes, no processo industrial para obtenção dos produtos cárneos processados, possivelmente resultaria em maior atividade antimicrobiana do mesmo contra S. Enteritidis / Abstract: Salmonella is one of the most important pathogens involved in foodborne outbreaks related to the consumption of ready-to-eat meat products. The contamination of these products can occur during processing operations like slicing, packaging or other handling procedures. The increase in demand for turkey breast based products has been verified worldwide since they combine the convenience of ready-to-eat-products with the fact that poultry meat is recognized as healthy by consumers. The presence of chemical preservatives in food products is another important factor that influences consumer¿s choice. The use of natural substances as antimicrobials, such as essential oils, and their incorporation in active packaging systems represents an interesting alternative to the use of chemical preservatives. The aim of this study was to develop and implement an edible coating incorporated with essential oil as a natural antimicrobial to turkey breast products and to assess its activity against Salmonella Enteritidis, in order to simulate an active packaging system. This study also aimed to evaluate the transfer of S. Enteritidis to turkey breast products during the slicing procedure, simulating a cross-contamination scenario. Pathogen behavior during sliced turkey breast products¿ shelf-life at different storage temperatures (10 e 25 °C) was also evaluated. Four different formulations of products (A, B, C and D) were studied in order to evaluate how the composition of these food matrices can influence the behavior of S. Enteritidis. Among the four essential oils tested in vitro, Thymus vulgaris (thyme) essential oil exhibited the lowest minimum inhibitory concentration (MIC) against the pathogen and it was selected to subsequent assays. Based on sensory analysis and antimicrobial screening tests, 0.8 % (v/w) of T. vulgaris essential oil was applied on the edible coatings to be tested on turkey breast meat products. S. Enteritidis could be transferred during the slicing procedure, from artificially contaminated slicer blades (~ 8 log CFU/blade) to all slices microbiologically analyzed (n = 20), of products previously coated and not coated with T. vulgaris essential oil. Statistical differences were detected regarding pathogen transfer in the different treatments (p ? 0.05). S. Enteritidis behavior on slices during storage was strongly dependent on temperature and products composition. Growth was verified in slices of all formulations and treatments during storage at 25 °C. Only one product was susceptible to pathogen growth at 10 °C (product A). In the slices of the other three products (B, C and D) a survival pattern was observed, with reductions of S. Enteritidis counts during shelf-life. T. vulgaris essential oil was effective against S. Enteritidis during storage of slices at 10 °C. Final tests indicated that the addition of T. vulgaris essential oil with the other ingredients, during the industrial manufacture process of turkey breast products, probably would result in higher antimicrobial activity against S. Enteritidis. / Mestrado / Ciência de Alimentos / Mestra em Ciência de Alimentos

As bactérias patogênicas são os principais agentes que contaminam alimentos e podem prejudicar a saúde humana. Para tentar combater e controlar a contaminação de alimentos investigam-se novos compostos que apresentam atividade antimicrobiana. O ramnolipídeo (RL) é um biossurfatante (BS) produzido por Pseudomonas spp. que apresenta elevada biodegradabilidade e, baixa toxicidade além de potencial antimicrobiano. O objetivo desse trabalho foi estudar a atividade antimicrobiana do RL frente às bactérias patogênicas Gram positivas, Bacillus cereus (ATCC 33018), Listeria monocytogenes (ATCC 19112), Staphylococcus aureus (ATCC 8095) e Gram negativas, Escherichia coli (EHEC) (ATCC 43895) e Salmonella enterica (ATCC 13076) além de contribuir na elucidação do mecanismo de ação destes compostos. Os testes de susceptibilidade ao RL foram realizados a partir da determinação da concentração inibitória mínima (CIM) e concentração bactericida mínima (CBM) utilizando a técnica de micro-diluição. O efeito do pH sobre a atividade antimicrobiana foi avaliado na faixa de pH 5 a 9. Para avaliação do mecanismo de ação foram realizados ensaios de permeabilidade celular, espectroscopia de infravermelho e hidrofobicidade celular. O RL apresentou atividade antimicrobiana para as bactérias B. cereus em CIM 19,5 &#956g/mL e CBM 39,1 &#956g/mL, e para L. monocytogenes CIM 156,2 &#956g/mL e CBM 312,5 &#956g/mL. Para B. cereus apresentou efeito bactericida a partir de 30 minutos na CBM, e para L. monocytogenes em 8 horas de incubação com o RL na CBM. As bactérias Gram negativas E. coli e S. enterica mostraram-se resistentes ao RL. O pH influenciou a ação antimicrobiana do RL sendo mais efetivo em pH mais ácidos. O tratamento com RL promoveu redução da hidrofobicidade da superfície celular das bactérias sensíveis. Os espectros infravermelhos evidenciaram alterações na composição química da membrana/parede celular principalmente para bactérias Gram positivas. A permeabilidade da membrana celular aumentou de acordo com o aumento da concentração de RL. A atividade antimicrobiana do RL foi evidenciada para as bactérias Gram positivas sendo mais sensíveis B. cereus e L. monocytogenes. Os resultados obtidos neste trabalho sugerem que o RL promove alterações na permeabilidade e composição química da membrana celular bacteriana sendo um agente potencial para controle de bactérias Gram positivas de importância alimentar. / Pathogenic bacteria are main agents that contaminate food and are harmful to human health. The search for new compounds to combat and control food pathogens is of increasing interest. Rhamnolipid (RL) is a biosurfactant (BS) typically produced by Pseudomonas spp., showing high biodegradability, low toxicity and antimicrobial activity. This study aimed to evaluate the antimicrobial activity of RL against the food pathogenic Gram positive bacteria Bacillus cereus (ATCC 33018), Listeria monocytogenes (ATCC 19112), Staphylococcus aureus (ATCC 8095) and Gram negative, Escherichia coli (EHEC) (ATCC 43895) and Salmonella enterica (ATCC 13076) and also contribute to the elucidation of RL mechanism of action. Susceptibility tests were performed by determination of the minimum inhibitory concentration (MIC) and minimal bactericidal concentration (MBC) using the broth microdilution method. The effect of pH on antimicrobial action was also investigated ranging from 5 to 9. Mechanism of action was studied using membrane permeability, infrared spectroscopy and cell hydrophobicity assays. The MIC value for B. cereus was 19.5 &#956g/mL and MBC was 39.1 &#956g/mL. L. monocytogenes was inhibited at concentration 156.2 &#956g/mL showing MBC of 312.5 &#956g/mL. B. cereus presented bactericidal effect after 30 minutes and for L. monocytogenes after 8 hours. The Gram-negative E. coli and S. enterica were resistant to RL. The pH influence antimicrobial activity of the RL showing decreasing MIC values at acidic conditions. Cell hydrophobicity was reduced by RL for the sensitive bacteria. Infrared spectroscopy showed that RL induced changes in chemical composition of cell membrane/ wall especially for the Gram positive bacteria. Cell permeability also increases as RL concentration increases. Antimicrobial activity of RL was evidenced for Gram positive bacteria and the most sensitive were B. cereus and L. monocytogenes. The results of this study suggest that rhamnolipid biosurfactant promotes changes in the permeability and membrane chemical composition showing potential to control foodborne Gram positive bacteria.

<p>U okviru ove disertacije uspe&scaron;no je sintetisano 14 novih protičnih jonskih tečnosti na bazi amina. Strukture su potvrđene snimanjem NMR i IC spektra. Izmerena je gustina svih jonskih tečnosti u temperaturnom opsegu&nbsp; T&nbsp; = (293,15&nbsp; &ndash;&nbsp; 353,15) K, kao i gustina njihovih vodenih rastvora u temperaturnom opsegu&nbsp; T&nbsp; = (293,15 &ndash; 313,15) K. Fizičko-hemijska karakterizacija je takođe propraćena merenjem viskoznosti i električne provodljivosti binarnih sme&scaron;a u celom rasponu&nbsp; molarnih frakcija i na različitim temperaturama (T&nbsp; = (293,15&nbsp; &ndash;&nbsp; 313,15) K). Na osnovu dobijenih rezultata utvrđeno je da sve protične jonske tečnosti imaju izražen structure making efekat. Ispitana je antimikrobna aktivnost protičnih jonskih tečnosti na bazi amina prema deset sojeva fungi: Trichoderma sp., Penicillium sp.(1-3), Cladoporium sp., Aspergillus sp. (1,2), A. flavus, F. graminearum i P. citrinium,&nbsp; osam sojeva bakterija:&nbsp; B. subtillis (6633), S. aureus (6538), E. faecalis (19433), K. rhizophila (9341),&nbsp; E. coli (11229), S. enteritidis (13076), P. mirabilis (12453), P. aeruginosa (15692) i dva soja kandida:&nbsp; C. albicans (ATCC 10231) i C. albicans (L) .&nbsp; Rezultati su pokazali da svih četrnaest jonskih tečnosti pokazuju antimikrobno delovanje. Za proučavane protične&nbsp; jonske tečnosti, antimikrobna aktivnost zavisi od dužine alkil lanca, pri tome pokazujući maksimalnu aktivnost sa anjonima koji sadrže 3 i 4 atoma ugljenika u nizu. Rezultati su pokazali da PILs pokazuju bolje antimikrobno dejstvo ako istovremeno sadrži i terminalno hlorovani C atom. Međutim, PILs pokazuju veću aktivnost ako je terminalni C atom monohalogeniran, jer monohalogenirani derivati poseduju veći afinitet za interakciju sa ćelijskom membranom. Najefikasnije PILs (ClA, 3-ClP, 4-ClB, Ac i For) su izabrane da se testiraju kao alternativni rastvarači umesto timola. Svih pet protičnih jonskih tečnosti uspe&scaron;no je testirano kao zelena alternativa za uobičajne organske rastvarače koji se koriste za či&scaron;ćenje i konzervaciju papirne povr&scaron;ine. Značajna antimikrobna svojstva primećena u ovoj doktorskoj tezi sugeriraju da proučavane protične jonske tečnosti mogu imati potencijalnu primenu u radu sa papirnim atifaktima, kao i pri njihovom či&scaron;ćenju i konzervaciji, zamenjujući na taj način primenu konvencionalnih rastvarača i organske supstance otrovne za ljude i životnu sredinu.</p> / <p>In this doctoral thesis the 14 amine based protic ionic liquids were newly synthesized. The validity of all structures were checked and confirmed by NMR and IR spectroscopy. Densities of pure ionic liquids and their diluted aqueous solutions have been measured over the whole composition range at selected temperatures from (293.3 to 353.15) K for pure ionic liquids, and from (293.15 to 313.15) K for diluted aqueous solutions. Physico-chemical&nbsp; characterization was also performed&nbsp; measuring the viscosity and electrical conductivity of binary mixtures in the entire range of mole fractions and at different temperatures (T&nbsp; = (293.15&nbsp; &ndash;&nbsp; 313.15) K). It was concluded that all investigated PILs have structure making tendency. Further, antimicrobial activity of amine based protic ionic liquids has been investigated using ten fungi strains: Trichoderma sp., Penicillium sp. (1-3), Cladoporium sp., Aspergillus sp. (1,2), A. flavus, F. graminearum i P. citrinium, eight ATCC bacterial strains: B. subtillis (6633), S. aureus (6538), E. faecalis (19433), K. rhizophila (9341), E. coli (11229), S. enteritidis (13076), P. mirabilis (12453), P. aeruginosa (15692) and two Candida&nbsp; strains:&nbsp; C. albicans (ATCC 10231) i C. albicans (L). The results indicated that all fourteen ionic liquids showed antimicrobial activity.&nbsp; For the studied PILs, the antimicrobial activity dependent on the alkyl chain length, showing maximal activity with anions containing 3 and 4 carbon atoms. The results revealed that the PILs show better antimicrobial activities if the longer alkyl chain is presented but in the same time contain chlorinated terminal C atom. However, PILs showed higer activity if the terminal C atom is monohalogenated, since the monohalogenated derivatives possess higher affinity to interact with cell membrane. The most effective PILs (ClA, 3-ClP, 4-ClB, Ac and For)&nbsp; were chosen to be tested as alternative solvents instead of thymol. All five PILs were successfully tested as the green alternatives to conventionally used organic solvents for paper cleaning and conservation. The significant antimicrobial properties observed in this doctoral thesis suggest that studied PILs may have potential applications in the paper art and artefact cleaning and conservation replacing thus, conventional solvents and organic substances that are toxic for humans and environment.</p>

La résistance aux antimicrobiens est un problème mondial majeur de plus en plus préoccupant. Le développement de nouveaux antimicrobiens classiques remplaçant les agents rendus inefficaces est certes l’une des stratégies les plus prometteuses visant à diminuer son ampleur. Mais cette solution demeure malheureusement momentanée. En fait tout agent antimicrobien sera tôt ou tard vaincu par la propagation des souches résistantes favorisée par le phénomène de pression de sélection exercée par l’agent lui-même. Il est donc nécessaire d’envisager toutes les alternatives possibles aux approches classiques, cela afin de diversifier l’arsenal thérapeutique antimicrobien.Les huiles essentielles utilisées par les plantes dans leur défense contre les pathogènes sont naturellement composées d’un mélange de constituants les rendant capables d’agir sur plusieurs cibles de l’organisme. De ce fait, même les pathogènes les plus résistants ne vont pouvoir échapper à leurs actions et le développement des résistances sera par conséquent limité. Dans le cadre de la présente thèse des plantes aromatiques libanaises sélectionnées en se basant surtout sur des indications ethno pharmacologiques ont été récoltées de différentes régions libanaises. Elles ont été par la suite hydrodistillées pour produire une vingtaine d’huiles essentielles qui ont été analysées par GC/MS et éventuellement par RMN, et évaluées contre une gamme de pathogènes responsables d’infections cutanées chez l’homme. L’analyse de la composition chimique des HEs a été marquée par l’isolement et la caractérisation pour la première fois du santolinoïdol, un sesquiterpène du type bisabolène qui a été retrouvé dans l’huile essentielle d’Achillea santolinoides subsp. wilhelmsii. Presque la moitié des HEs ont été actives sur au moins un des pathogènes. L’origine de l’activité des huiles les plus actives, l’effet de leurs associations avec des antimicrobiens commerciaux ainsi que leurs cytotoxicités ont été également investigués. / Antimicrobial resistance is a major global problem and a growing concern. The development of new conventional antimicrobial agents replacing inefficient ones is certainly one of the most promising strategies aiming to reduce its extent. But this solution remains unfortunately momentary. In fact, any antimicrobial agent will eventually be defeated by the spread of resistant strains favored by the selection pressure phenomenon exerted by the agent itself. It is therefore necessary to consider all possible alternatives to conventional approaches in order to diversify the antimicrobial arsenal.Essential oils used by plants in their defense against pathogens are naturally composed of a mixture of components making them able to act on several targets of the organism. Thus, even the most resistant pathogens will not be able to escape their actions and the development of resistance will be therefore limited.In this thesis, Lebanese aromatic plants selected based mainly on ethnopharmacological indications were collected from various Lebanese regions. They were subsequently hydrodistillated producing twenty essential oils analyzed by GC / MS and eventually by NMR, and assessed against a range of pathogens that cause skin infections in humans. The analysis of the chemical composition of the EOs was marked by the isolation and characterization for the first time of santolinoïdol, a bisabolene type sesquiterpene that was found in the essential oil of Achillea santolinoides subsp. wilhelmsii. Almost half of the EOs have been active on at least one pathogen. The origin of the activity of the most active oils, the effect of their associations with commercial antimicrobials and their cytotoxicities were also investigated.

Diplomová práce se zabývá vlivem uhlíkatého plniva na životaschopnost a produkci extracelulárních látek vybrané bakterie Bacillus subtilis (CCM 1999) a kvasinky Yarrowia lipolytica (CCY 29-26-52). Antimikrobiální aktivita těchto částic, přítomných v kultivačním mediu, byla sledována pomocí následujících parametrů: růst daného mikroorganismu, produkce extracelulárních proteinů a v poslední řadě byla monitorována produkce extracelulárních polymerních substancí, které mají úzkou souvislost s tvorbou biofilmu. Suspenze materiálů (0,135 mg/mL) byly připraveny ve dvou rozdílných kultivačních mediích; tzn. živné medium s obsahem glukózy pro Bacillus subtilis a bazální medium s přídavkem Tweenu 80 pro Yarrowia lipolytica, a media byla inokulována příslušným typem mikroorganismu. Experimenty probíhaly po dobu 6 dnů při rychlosti třepání 160 rpm a teplotě 30 °C pro Bacillus subtilis a 28 °C pro Yarrowia lipolytica. Testovány byly celkem tři typy uhlíkatého nanomateriálu, získané z Katedry anorganické chemie, Vysoké školy chemicko-technologické v Praze. Tyto materiály specifikované jako materiál “A”, “B” a “C” se navzájem lišily velikostí částic a stupněm oxidace. Na základě skríningových studií byla vybrána koncentrace testovaného materiálu 0,135 mg/mL a rychlost třepání 160 rpm. Metodou měření optické hustoty vzorku při 600 nm byly sestaveny a porovnány růstové křivky obou mikroorganismů v přítomnosti testovaných nanočástic po dobu 5 dní. Tímto způsobem bylo zjištěno, že přítomnost nanočástic v mediu nemá velký vliv na růst zkoumaného mikroorganismu. Tato metoda, je však pouze orientační, protože se nevyhneme chybě díky přítomnosti mrtvých buněk. Dále byla testována produkce celkových a extracelulárních proteinů daným mikroorganismem v přítomnosti testovaných nanočástic. Nebyla však pozorována výrazná odchylka hodnot od hodnot kontrolního vzorku, který neobsahoval testovaný materiál. Na základě metod počítání kolonií (Bacillus subtilis) a buněk (Yarrowia lipolytica) byly určeny ztráty životaschopnosti mikroorganismu ve 3 časech (6, 48 a 144 hodin); v kratším časovém intervalu byl růst spíše podporován. Dále byla monitorována produkce extracelulárních polymerních substancí (EPS), tedy proteinů, redukujících substancí a polysacharidů. Tyto látky byly vylučovány daným mikroorganismem do prostředí v průběhu 24 hodin. Bacillus subtilis produkoval EPS ve větší míře než Yarrowia lipolytica. Předpokládáme, že produkce EPS by mohla souviset s tvorbou biofilmu, který chrání buňky před toxicitou nanočástic.

Present diploma thesis is focused on natural antimicrobial substances. The aim of the experimental part of thesis was testing of antimicrobial activity of selected natural substances in free and encapsulated forms, preparation of particles and monitoring of their long-term stability. Various teas extracts (aqueous, ethanoic and DMSO) and aqueous herbs extracts were used as plant antimicrobial substances. Liposomes and chitosan particles with encapsulated aqueous teas extracts were prepared by ultrasonication. Encapsulation efficiency, size, stability and antioxidant activity for all particles were specified. Consequently, within a two months period, long-term stability for these particles was monitored. Spectrophotometric method for determining total polyphenols was used for evaluating encapsulation efficiency as well as for monitoring of long-term stability. The DLS method was used for determination of size and stability of particles. Antioxidation activity was monitored by using spectrophotometric method using ABTS. For testing antimicrobial activity of various types of teas, herbs extracts and their mixtures and combianitons with lysozyme and antibiotics, broth dilution methods were used while using both Gram positive and Gram negative bacterial test strains. Antimicrobial activity of prepared particles was also monitored, the antimicrobial activity was especially significant with the chitosan particles. Suitable combinations of natural substances whatever with antimicrobial or antioxidation effect either in free or encapsulated form could be used for applications for example in cosmetics industry, food processing or in medicine.

M. Tech. (Department of Chemistry, Faculty of Applied and Computer Sciences), Vaal University of Technology. / Electrospinning is a method that has gained more attention due to its capability in spinning a wide variety of polymeric fibers and nanoparticles embedded in polymer fibers. Polymer blending has been considered the most appropriate way for creating new materials with fused properties which improve poor chemical, mechanical, thermal and dynamic mechanical properties of each polymer. Hence, in this study, electrospinning technique was used to fabricate polyacrylonitrile (PAN) nanofibers at concentrations of (10, 12 and 14 wt%) and cellulose acetate (CA) nanofibers at concentrations of (14, 16 and 18 wt%). 10wt% of PAN and 16 wt% of CA were blended together and the optimum blend ratio was found to be 80/20 PAN/CA. TiO2 nanoparticles (0.2 and 0.4 wt%) were incorporated into CA nanofibers and (1, 2 and 3 wt%) were incorporated into PAN and PAN/CA blended polymers, respectively. Applied voltages of 20, 22 and 24 kV were varied at a spinning distance of 15cm and the optimum voltage for the fabrication of composite was 22 kV. The sol-gel method was used to synthesise the TiO2 nanoparticles at different calcination temperatures of 400, 500 and 600 ºC. The fabricated composite nanofibers were tested for antibacterial and photocatalytic activities. The synthesised nanomaterials were characterized using SEM, TEM, EDX, UV-Vis, PL, FTIR spectroscopy, XRD and TGA. The absorption and emission spectra illustrated the formation of TiO2 nanoparticles and the increase in absorption band edges. TEM showed the spherical morphology of the nanoparticles with average diameter of 12.2 nm for nanoparticles calcined at 500 ºC. SEM illustrated the diameter and morphology of the nanofibers and composites with the average diameter of 220, 338, 181, 250, 538, 294 nm for PAN, CA, PAN-TiO2, CA-TiO2, PAN/CA and PAN/CA-TiO2, respectively. XRD revealed anatase phase as the dominant crystalline phase of the synthesised nanoparticles. FTIR spectroscopy and EDX signified that the formation of composite nanofibers and the presence of TiO2 nanoparticles corresponded to the Ti-O stretching and Ti-O-Ti bands on the FTIR spectra. The antimicrobial activity of the composite nanofibers were tested against E. coli, S. aureus and C. albicans microorganisms. The photocatalytic activity of the nanomaterials was tested using methyl orange dye. PAN/CA-TiO2 composite nanofibers revealed the greatest antibacterial activity against selected microorganisms as compared to the other nanocomposites. PAN/CA-TiO2 nanocomposite (44%) showed greater rate of photodegradation of methyl orange than PAN-TiO2 nanofibers (28%) and TiO2 nanoparticles (12%) under visible light irradiation.