O presente trabalho tem como proposição investigar a hipótese de que a diminuição da tensão superficial do líquido favorece o comportamento reológico de suspensões concentradas, uma vez que as partículas estão mais próximas e a contribuição da força de capilaridade possivelmente aumenta. Como suspensões concentradas tendem ao comportamento viscoelástico, investigou-se métodos reológicos propícios para analisar possíveis consequências devido à alteração da tensão superficial da água. Foram utilizados dois carbonatos de cálcio equivalentes (P1 e P5) que se diferenciam pela distribuição granulométrica. Utilizou-se dois dispersantes de mercado e etilenoglicol como modificadores de tensão superficial. A análise de superfície pelo ensaio de potencial zeta, revelou carga superficial positiva. Embora os pós sejam equivalentes, a mobilidade eletroforética do P5 é menor. Verificou-se que o íon cálcio é determinante do potencial. Verificou-se contribuição eletrostática apenas dos dispersantes. O ensaio de gota pendente constatou diminuição da tensão superficial da água com os três aditivos. Ensaios de ascensão capilar pelo método de Washburn revelaram maiores ângulos de contato para ensaios com aditivos, sendo menor para P5. Avaliou-se geometrias e métodos reológicos a fim de selecionar bom conjunto para medidas de viscoelasticidade, sendo oscilatório de tensão e geometria vane escolhidos. O acréscimo dos dispersantes resultou em menores valores de G\' e tensões de escoamento, enquanto o etilenoglicol resultou em maiores. Não há relação clara entre valores calculados de ângulo de contato e mobilidade eletroforética. Os ensaio reológicos e de mobilidade relacionaram-se apenas para os dispersantes. A hipótese foi verificada pelos valores de tensão de escoamento e IPS. Para os dispersantes, foi possível observar comportamento próximo à hipótese, quanto menor foi a tensão superficial do líquido, menores valores de tensão para menores IPS. Explorou-se também a hipótese pelos ensaios de ângulo de contato, observou-se novamente curva próxima a hipótese. Embora necessidade de mais ensaios, o presente trabalho contribuiu para metodologia de exploração de características de superfície, dispersão e reológicas de suspensões concentradas. / The proposition of the present work is to investigate the hypothesis that the reduction of the surface tension of the liquid favors the rheological behavior of concentrated suspensions, since the particles are closer and the contribution of capillarity forces possibly increases. As concentrated suspensions tend to viscoelastic behavior, rheological methods were examined to analyze possible consequences due to changes of surface tension of the water. Two equivalent calcium carbonates (P1 and P5) were used and differ by particle-size distribution. Two market dispersants and ethylene glycol were used as surfactant. Surface analysis by the zeta potential test revealed positive surface charge and, although the powders are equivalent, the electrophoretic mobility of P5 is lower. The reults show that calcium is potential determining ion. Electrostatic contribution was found only for the dispersants. Pendant drop test showed a decrease in the surface tension of the water with the three additives. Washburn capillary rise technique estimated greater contact angles for tests with additives and lower ones for P5. Rheological geometries and methods were evaluated in order to select good set for viscoelasticity measurements, choosing oscillatory stress sweep and vane geometry. The addition of the dispersants resulted in lower values G \'and yield stress, while ethylene glycol resulted in higher values. There is no clear relation between contact angle and electrophoretic mobility. The rheological and eletrophoretic mobility showed relation only for dispersants. The hypothesis was verified by the values yield stress and IPS. For the dispersants, it was possible to observe behavior close to the hypothesis, the lower the surface tension of the liquid, the lower the yield stress for lower IPS. The hypothesis was also explored by the contact angle tests, a curve next to the hypothesis was observed again. Although the need for more tests, the present work contributed to the methodology of exploration of surface characteristics, dispersion and rheological characteristics of concentrated suspensions.

L'objectif principal de cette thèse a été d'étudier et de développer les méthodes électrophorétiques pour la détermination de la charge effective de petits ions, de (bio)macromolécules et de nanoparticules. En effet, la charge effective est un paramètre physico-chimique qui contrôle les interactions électrostatiques et qui permet d'accéder aux taux de condensation des contre-ions dans le cas de polyélectrolytes. Dans une première partie, différents modèles sur la mobilité électrophorétique (Nernst-Einstein, O'Brien-White-Ohshima, Yoon-Kim) ont été comparés pour la détermination de la charge effective à partir des valeurs expérimentales de mobilité électrophorétique et de rayon hydrodynamique. Trois autres méthodes expérimentales basées sur la sensibilité de détection UV en mode indirect, sur la sensibilité de détection en conductimétrie et sur la longueur des zones isotachophorétiques ont été étudiées. Ces méthodes ont été appliquées en particulier à la détermination de la charge effective de dendrimères greffés de la lysine et de polymères utilisés en délivrance de principe actif.Une étude du comportement électrophorétique en fonction de la force ionique nous a mené à proposer une représentation graphique, appelée « slope-plot », permettant de distinguer les solutés en fonction de leur nature (petits ions, polyélectrolytes, nanoparticules). Cette représentation peut s'avérer très utile pour l'optimisation des séparations en électrophorèse capillaire en fonction de la force ionique. / The main objective of this thesis was to study and develop electrophoretic methods for effective charge determination of small ions, (bio)macromolecules and nanoparticles. Effective charge is a physical parameter that controls the electrostatic interactions and allows for the determination of condensed counter-ion fraction in the case of polyelectrolytes. In a first part, different models of electrophoretic mobility (Nernst-Einstein, O'Brien-White-Ohshima, Yoon-Kim) have been compared for effective charge determination from experimental values of electrophoretic mobility and hydrodynamic radius. Three other experimental methods based on the sensitivity of UV detection in indirect mode and in conductivity detection, or on the length of the isotachophoretic zones, were studied. These methods were applied to effective charge determination of dendrigraft poly-L-lysines and on drug delivery polymeric systems. A study of the ionic strength dependence of the electrophoretic mobility leads us to propose a graphical representation, called the slope-plot, allowing for the distinction between solutes according to their nature (small ions, polyelectrolytes, nanopaticles). The slop-plot can also be used for the optimization of electrophoretic separations according to the ionic strength.

Dissertation (Ph.D.)--University of South Florida, 2009. / Title from PDF of title page. Document formatted into pages; contains 86 pages. Includes vita. Includes bibliographical references.

Dissertation (Ph.D.)--University of South Florida, 2009. / Includes vita. Includes bibliographical references. Also available online.

Orientadores: Maria Isabel Nogueira Cano, Carlos Henrique Inacio Ramos / Tese (doutorado) - Universidade Estadual de Campinas, Instituto de Biologia / Made available in DSpace on 2018-08-08T19:46:21Z (GMT). No. of bitstreams: 1 Lira_CristinaBragadeBrito_D.pdf: 9068778 bytes, checksum: 7561201fa37b1109dc7803fb6173aad0 (MD5) Previous issue date: 2007 / Resumo: A leishmaniose é uma doença parasitária que foi considerada pela Organização Mundial de Saúde como uma doença de Categoria 1, pois para a mesma não existem formas de controle, diagnóstico ou terapia eficientes. Os parasitas do gênero Leishmania (família Trypanosomatidae) são agentes etiológicos da leishmaniose e possuem cromossomos lineares com extremidades teloméricas. Os telômeros são complexos nucleoproteicos essenciais para manuntenção da estabilidade do genoma e viabilidade celular. Devido à importância das proteínas teloméricas na manutenção dessas estruturas, elas têm sido considerados bons alvos para a terapia anticâncer e antiparasitária. Sendo assim, o objetivo deste trabalho foi identificar proteínas que se associam aos telômeros de Leishmania amazonensis. Três metodologias diferentes foram utilizadas para antigir este objetivo: (i) purificação de proteínas teloméricas a partir de extratos de L. amazonensis, (ii) busca no banco de dados de Leishmania por proteínas homólogas que apresentassem domínios de ligação ao DNA do tipo Myb, e (iii) seleção genética em levedura (sistema mono-híbrido). A purificação de proteínas teloméricas a partir de extratos nucleares de L. amazonensis resultou no isolamento da proteína LaRbp38. Ensaios in vitro de interação proteína-DNA e ensaios in vivo (imunoprecipitação de cromatina) comprovaram a interação de LaRbp38 com DNA telomérico, DNA rico em GT e DNA do cinetoplasto. LaRbp38 pode ser considerada um bom alvo para a terapia antiparasitária pois é uma proteína exclusiva de tripanosomatídeos e seu homólogo em T. brucei é essencial para a sobrevivência do parasita. Para dar início a experimentos de caracterização da estrutura dessa proteína, LaRbp38 foi expressa em bactérias. Como a proteína permaneceu na fração insolúvel, experimentos preliminares de renovelamento foram feitos mostrando que a proteína necessita da presença de DNA, ou de moléculas análogas, para se renovelar in vitro. As buscas no banco de dados de L. major resultaram no descobrimento de uma lista de proteínas que apresentam domínio de ligação ao DNA do tipo Myb. Uma proteína foi escolhida e sua homóloga em L. amazonensis foi denominada LaTBP1. O domínio Myb de LaTBP1 se localiza na porção central da proteína e apresenta alta similaridade de seqüência com domínios Myb encontrados em fatores de transcrição to tipo TFIIIB, proto-oncogene c-MYB e membros da família de proteínas teloméricas Rap1p. Ensaios de interação proteína-DNA e de imunoprecipitação de cromatina confirmaram que LaTBP1 interage tanto com o DNA telomérico quanto com DNAs ricos em GT. Prefências por estes dois tipos de DNAs são apresentadas pelas proteínas teloméricas Rap1, Taz1 e TEBP1, descritas em leveduras e eucariotos superiores. A utilização do sistema mono-híbrido para identificação de proteínas teloméricas de Leishmania resultou em uma lista de proteínas candidatas. A possível função telomérica das mesmas foi inferida com base na homologia de seqüência com proteínas já descritas e em testes de interação proteína-DNA in vitro utilizando-se extratos das leveduras recombinantes. Apesar desses resultados serem promissores, análises mais detalhadas são necessárias para confirmar estas interações. Concluindo, as três metodologias se mostraram eficazes para a identificação de proteínas teloméricas e a utilização das mesmas resultou na identificação de diversas proteínas teloméricas, algumas delas ainda hipotéticas / Abstract: Leishmaniasis is a parasitic disease that was classified by World Health Organization as a Category 1 disease because there are no effective control programs or therapeutics to this disease. Parasites from the Leishmania genus are the ethiologic agents of leishmaniasis and they possess linear chromosomes with telomeric ends. Telomeres are nucleoprotein specialized nucleoprotein complexes essential for maintaining chromosomal stability and cell viability. Since telomeric proteins are essential for the maintenance of telomeres, they could be considered good targets for anticancer and antiparasitic drugs. Therefore, the goal of this work was to identify Leishmania amazonensis proteins that interact with the telomeric DNA. Three methodologies were used the achive this goal: (i) purification of telomeric proteins from nuclear extracts of L. amazonensis, (ii) Leishmania genome database mining for protein containing a Myb-like domain, and (iii) use of One-hybrid system (Clontech). The search for telomeric proteins in nuclear extracts of L. amazonensis resulted in the identification of LaRbp38. EMSA and immunoprecipitation assays were used to attest LaRbp38 binding to telomeric, GT-rich and kinetoplast DNAs, both in vitro and in vivo. LaRbp38 could be considered a good drug target for antiparasitic therapy since it is exclusive of trypanosomatids and itshomologue in T. brucei is essential for parasite survival. In order to characterize structurally this protein, LaRbp38 was expressed in bacteria. The protein was present in the insoluble fraction of the bacterial lysate. Therefore, preliminary experiments of refolding were done. LaRbp38 seems to need DNA, or analog molecules, in order to correctly refold in vitro. Data-mining in the L. major genome resulted on a list of proteins bearing a Myb-like domain. One protein was chosen and its L. amazonensis homologue was termed LaTBP1. LaTBP1 Myb-like domain is centrally localized and shares sequence similarities with Myb-like domains found on transcription factors TFIIIB and c-MYB, and with the RAP1 telomeric proteins. Competition and chromatin immunoprecipitation assays confirmed the specificity of LaTBP1 for telomeric and GT-rich DNAs. This binding specifities are also found on the telomeric proteins Rap1, Taz1 and TEBP1, described in yeast and higher eukaryotes. A list of proteins with a putative telomeric function was generated after the use of the Onehybrid system. Sequence analysis (search for homologues in other organisms) and EMSA, done with protein extract of recombinant yeast, were used to infer a telomeric function for the proteins found by this methodology. Although the results are encouraging, detailed analysis are necessary to validate the interactions. In conclusion, the three methodologies were usefull for the identification of telomeric proteins. This work resulted in the identification of several telomeric candidate proteins / Doutorado / Genetica de Microorganismos / Doutor em Genetica e Biologia Molecular

No description available.

No / The electrophoretic mobilities of three beta-blocker drug practolol, timolol and propranolol, have been measured in electrolyte systems with mixed binary and ternary water-methanol-ethanol solvents with acetic acid/sodium acetate as buffer using capillary electrophoresis. The highest mobilities for the analytes studied have been observed in pure aqueous. the lowest values in ethanolic buffers The measured electrophoretic mobilities have been used to evaluate the accuracy of a mathematical model based on a mixture response surface method that expresses the mobility as a function of the solvent composition. Mean percentage error (MPE) has been computed considering experimental and calculated mobilities as an accuracy criterion. The obtained MPE for practolol, timolol and propranolol in the binary mixtures are between 0.9 and 2.6%, in the ternary water-methanol-ethanol solvent system the MPE was about 2.7%. The MPE values resulting from the proposed equation lie within the experimental relative standard deviation values and can he considered as an acceptable error.

This study was conducted to determine how the structure and surface chemistry of bulk CaCO3 differs from that of nanometer-sized CaCO3 and then to determine rate, extent and mechanisms of As adsorption on various synthetic CaCO3 materials. Additionally, we sought to devise a chemical CaCO3 precipitate that approximates biogenic CaCO3. The bulk CaCO3 precipitation was performed by using a solution that was highly oversaturated so that large CaCO3 precipitates rapidly form. Two different methods were employed for the synthesis of nanometer size CaCO3 i) an in situ deposition technique and ii) an interfacial reaction (water in oil emulsion). Mineral characterization of all CaCO3 precipitates was done with Nitrogen Porosimetry (Brunauer Emmett Teller method), particle size analysis, X-ray diffraction and Fourier Transform Infrared/ Fourier Transform Raman spectroscopy. The principal objective of the research was to assess the overall reactivity of As(III) and As(V) with different synthetic CaCO3 minerals. This was accomplished by i) running adsorption isotherms (varying As concentration), ii) measuring pH envelopes (varying pH at a fixed concentration) and iii) kinetic experiments (varying reaction time). Also, electrophoretic mobility experiments were performed in the presence of As(III) and As(V), and these studies revealed that As(III) forms stronger inner-sphere complexes with CaCO3 than As(V). Also, it was found that nanometer-sized CaCO3 prepared via deposition formed stronger inner-sphere complexes with As oxyanions (q = 5.26 µmol/m2) compared to either nano-sized CaCO3 from interfacial reactions (q = 4.51 µmol/m2) or bulk CaCO3 (q = 4.39 µmol/m2).<p> The PEG-based nano CaCO3 prepared by an in-situ deposition technique presents a novel and readily available synthesis route that can be used as proxy for the biogenic CaCO3 known to be present in many different environmental conditions. The results of this study suggest that CaCO3 can be used as a sorbent for As in groundwater.

This study was conducted to determine how the structure and surface chemistry of bulk CaCO3 differs from that of nanometer-sized CaCO3 and then to determine rate, extent and mechanisms of As adsorption on various synthetic CaCO3 materials. Additionally, we sought to devise a chemical CaCO3 precipitate that approximates biogenic CaCO3. The bulk CaCO3 precipitation was performed by using a solution that was highly oversaturated so that large CaCO3 precipitates rapidly form. Two different methods were employed for the synthesis of nanometer size CaCO3 i) an in situ deposition technique and ii) an interfacial reaction (water in oil emulsion). Mineral characterization of all CaCO3 precipitates was done with Nitrogen Porosimetry (Brunauer Emmett Teller method), particle size analysis, X-ray diffraction and Fourier Transform Infrared/ Fourier Transform Raman spectroscopy. The principal objective of the research was to assess the overall reactivity of As(III) and As(V) with different synthetic CaCO3 minerals. This was accomplished by i) running adsorption isotherms (varying As concentration), ii) measuring pH envelopes (varying pH at a fixed concentration) and iii) kinetic experiments (varying reaction time). Also, electrophoretic mobility experiments were performed in the presence of As(III) and As(V), and these studies revealed that As(III) forms stronger inner-sphere complexes with CaCO3 than As(V). Also, it was found that nanometer-sized CaCO3 prepared via deposition formed stronger inner-sphere complexes with As oxyanions (q = 5.26 µmol/m2) compared to either nano-sized CaCO3 from interfacial reactions (q = 4.51 µmol/m2) or bulk CaCO3 (q = 4.39 µmol/m2).<p> The PEG-based nano CaCO3 prepared by an in-situ deposition technique presents a novel and readily available synthesis route that can be used as proxy for the biogenic CaCO3 known to be present in many different environmental conditions. The results of this study suggest that CaCO3 can be used as a sorbent for As in groundwater.

Les bactériocines de classe IIa présentent une activité antimicrobienne résultant d'un mécanisme d'action ciblant les membranes des bactéries à Gram positif. Cette activité est modulée par différentes caractéristiques des surfaces bactériennes. Les propriétés physico-chimiques de surface de dix-huit souches bactériennes ont été déterminées afin d'étudier le lien entre ces propriétés et les phénotypes de résistance/sensibilité à Cbn BM1. Les résultats obtenus indiquent une grande diversité des propriétés physico-chimiques des surfaces analysées, sans cependant permettre d’établir un lien entre celles-ci et le phénotype de sensibilité/résistance à CbnBM1. Les mécanismes d'action de Cbn BM1 ont ensuite été étudiés sur Carnobacterium maltaromaticum DSM20730 et Listeria monocytogenes EGDe. L'atteinte de l'intégrité physique des membranes plasmiques par l'action de Cbn BM1 montre une hétérogénéité de réponse des populations bactériennes. Ce résultat a été confirmé par microscopie de force atomique in vivo à haute résolution. L'interaction de Cbn BM1 avec les membranes a été mise en évidence par mesure de l'anisotropie de fluorescence. Cette approche a révélé que Cbn BM1 présente des degrés de pénétration différents dans la membrane de C. maltaromaticum DSM20730 par rapport à L. monocytogenes EGDe. L'action de Cbn BM1 conduit cependant, pour les deux souches, à la modification de la force protomotrice membranaire. Ces différentes approches retenues pour l'étude des mécanismes d'action ont révélé que C. maltaromaticum DSM20730 et L. monocytogenes EGDe présentent une sensibilité à Cbn BM1 uniquement lorsque les cellules sont en phase exponentielle de croissance. / The antimicrobial activity of class IIa bacteriocins toward Gram positive bacteria relies on their membrane targeting mechanisms of action. These mechanisms are modulated by the bacterial surface properties. The physico-chemical surface properties of eighteen bacterial strains were determined to link these properties to the resistance/sensitivity to Cbn BM1 of the bacterial strains. In this way, two approaches were undertaken : the microbial adhesion to solvents and electrophoretic mobility measurements. The results show a large diversity of the determined properties among the strains but without establishing a direct link between the surface properties and the resistance/sensitivity phenotypes. Mechanisms of action of the bacteriocin Cbn BM1 on Carnobacterium maltaromaticum DSM20730 and Listeria monocytogenes EGDe were determined. Syto9® and propidium iodide allowed to show the heterogeneity of the bacterial populations toward the alteration of the membrane integrity. The interaction of Cbn BM1 with the bacterial membrane was studied by monitoring the fluorescence anisotropy of DPH and TMA-DPH. The results highlight a difference between the mechanism of action of Cbn BM1 on C. maltaromaticum DSM20730 and on L. monocytogenes EGDe. However, a treatment by Cbn BM1 leads to a perturbation of the component of the proton-motive force of the membrane for both strains. These approaches revealed that these bacterial strains exhibit a sensitivity to Cbn BM1 only when treated in log growth phase. Modification of nano-mechanical properties of C. maltaromaticum DSM20730 after a treatment by Cbn BM1 were assessed by an atomic force microscopy approach.