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Another family of organic anion transporters, the multi-drug resistant-associated-protein (MRP) is another subject of my study. Reverse transcription PCR and immunohistochernistry showed that MRP1 is expressed by the basal cells. MRP1 was cloned from the rat epididymis and expressed in HEK cells with a view to delineate its functions. HEK cells expressed with MRP1 showed little accumulation of calcein-AM compared to cells not expressed with the protein, or cells expressed with the protein but treated with MK571, an inhibitor of MRP1. This ability of MRP1 to eliminate calcein-AM from the cells was competitively reduced by verapamil, omeprazole, lonidamine or methotrexate, all are pharmacological agents commonly used for diverse clinical conditions. These results indicate that MRP1 is a multi-specific drug transporter. Freshly isolated basal cells also excluded verapamil and this effect was blocked by MK571. / In summary, my work has focused on the expression, distribution and functions of two major organic anion transporters, OAT1 and MRP1 thought to be the molecular entities involved in the transport of organic substances across the epididymal tubule. This membrane- and cell-specific placement of the two organic anion transporters that have different substrate specificity and mediate counter flows of substrates offer a mechanism whereby organic substance entry into the epididymis can be 'filtered'. Having low substrate specificity, OAT1 indiscriminately brings into the cells a diverse spectrum of organic anions, some of which might be useful organic substances that nourish spermatozoa, while others could be sperm toxicants. By virtue of the drug-specific MRP1, basal cells expel some of these toxicants from the epididymis. In this way, basal cells play a protective role. They prevent access of foreign substances to spermatozoa stored in the epididymis. This hypothesis is speculative albeit an attractive one. Further work is required to lay the experimental groundwork for this hypothesis. / The functions of OAT1 in the rat epididymis were studied by measuring the secretion (basal-to-apical) of PAH by epididymal epithelia in culture. The transport exhibited non-genomic up-regulation and down-regulation by PKA and PKC, respectively. These short-term regulations are not unlike those reported for the kidney OAT1 and are thought mediated through agonists-stimulated MAPK, PLA2 or COX-1 pathway. Since COX-1 has previously been shown to be exclusively present in the basal cells, it is conceivable that organic anion transport by the principal cells is regulated by the basal cells in the manner that principal cell Cl- secretion is regulated by the basal cells. / The nature of my work is to study the way by which organic substances cross the blood-epididymis barrier. My work is inspired by the anatomical and functional analogies between the epididymis and the kidney tubule. In the latter, elimination of toxic metabolites and foreign substances are known to occur through arrays of membrane proteins uniquely adapted to transport cationic and anionic organic substances across membranes. My work is to unravel the role of organic anion transporters in the epididymis using a combination of molecular biology and cellular physiology techniques carried out on cultured epithelia as well as on freshly isolated cells from the rat epididymis. / Using RT-PCR and Western Blot analysis, I identified the expression of organic anion transporter genes, OAT1, OAT2 and OAT3 mRNA in the rat epididymis. Immunohistochemistry revealed that OAT1 protein is present in the basal side of the principal cells of both the corpus and cauda epididymidis. OAT3 is seen in the lamina propria and blood vessels but not on the principal or other epithelial cells. Immunohistochemistry, however, failed to detect OAT2 protein. OAT1 was cloned from the rat cauda epididymidis using standard cloning techniques and verified by gene sequencing. Functional studies of the cloned protein expressed in human embryonic kidney (HEK) cells revealed similarities between the rat epididymal OAT1 and kidney OAT1 in that both transporters take up para-aminohippurate (PAH) into cells with similar kinetics and efficacy. Rat epididymal OAT1 was able to take up the anti-fertility drug, lonidamine. / Leung Chi Ting Gideon. / "October 2006." / Adviser: P. Y. D. Wong. / Source: Dissertation Abstracts International, Volume: 68-09, Section: B, page: 5678. / Thesis (Ph.D.)--Chinese University of Hong Kong, 2006. / Includes bibliographical references (p. 122-143). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Electronic reproduction. [Ann Arbor, MI] : ProQuest Information and Learning, [200-] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Abstracts in English and Chinese. / School code: 1307.

Anions have a wide range of importance both in chemical, as well as biological, systems; thus, the design and synthesis of novel receptors with the ability to selectively recognize or bind a specific class of anions is a rapidly developing field of supramolecular chemistry. A series of novel, acyclic pyrrole-based anion receptors will be presented. These systems, which are based on pyridine 2,6-dicarboxamides, bind nitrite and carboxylate anions with good selectivity in dichloroethane solution and are also capable of binding cyanide anions weakly. Control systems, incorporating a benzene-1,3-dicarboxamide spacer, or those wherein the connectivity of the amide linkage is "reversed," either failed to act as effective anion receptors or displayed very different selectivities. Such observations provide support for the notion that small perturbations in the structure of these receptors can lead to drastic changes in their anionbinding properties. Furthermore, efforts have been made to attach macrocyclic phosphate-binding receptors developed in the Sessler Group to cellulose solid supports. The idea is that these macrocycles, once bound to cellulose, will be capable of extracting phosphate from solutions. Studies on the macrocyclic loading level and extraction abilities of the receptors are underway, and will be presented herein. / text

Made available in DSpace on 2014-06-11T19:22:55Z (GMT). No. of bitstreams: 0 Previous issue date: 2010-01-22Bitstream added on 2014-06-13T19:47:18Z : No. of bitstreams: 1 silva_vct_me_sjrp.pdf: 648221 bytes, checksum: 471be77c5cc136374f3cf0a74fb31395 (MD5) / A ligação do O2 à Hemoglobina (Hb) é um processo regulado por interações alostéricas. Os íons Inositol hexafosfato (IHP) e cloreto, e prótons, são efetores alostéricos que estabilizam a Hb na estrutura quaternária T, diminuindo a afinidade da proteina pelo O2 em meio alcalino. A ligação de IHP também altera a cooperatividade de algumas hemoglobinas como, por exemplo a hemoglobina desArg, uma variante da HbAo da qual o resíduo arginina 141 das cadeia alfas foram retirados enzimaticamente. Estas argininas são prováveis sitios de interação com ions cloreto. Neste trabalho investigou-se a influência que a ligação de ânions Cl - e/ou IHP à Hb nas propriedades funcionais e conformacionais da proteína. Foi determinado o efeito destas interações na afinidade e cooperatividade de ligação de oxigênio à proteína, na liberação de prótons com a oxigenação ( efeito Bohr ), e na conformação da proteína. O papel de resíduos específicos nestes efeitos foi investigado comparando resultados obtidos com HbAo, Hb-desArg, e Hb Chesapeake (R92L), um mutante natural na qual a arginina 92 esta substituída por leucina. Os resultados mostraram que a retirada do resíduo C-terminal da cadeia alfa, arginina 141, suprimiu a cooperatividade da hemoglobina, diminui o número de prótons liberados durante a oxigenação, e que esta liberação de prótons foi a mesma na presença ou ausência de íons cloreto. Ainda, a retirada deste resíduo não interferiu no efeito Bohr dependente de IHP. Os resultados mostraram que a saturação de oxigênio ligados à HbA0 na presença de IHP não induziu a mudança na estrutura quaternária observada na ausência deste ânion, e na sua presença em meio com NaCl. Apesar disto, a cooperatividade da ligação é mantida. A arg-141α tem papel fundamental na estabilização pelo IHP da estrutura quaternária... / The hemoglobin (Hb) oxygenation is a molecular process regulated by allosteric interactions. Inositol hexaphosphate (IHP), chloride and protons are heterotropic effectors that stabilizes the deoxy state, the quaternary structure T, decreasing O2 affinity in alkaline solution. IHP binding also modifies cooperativity of several Hbs, such as desArg Hemoglobin, a variant form of HbA which is obtained by enzymatic cleavage of the two Arg-141alfa residues. This residue has been identified to be a Cl - binding site on Hb. This work follows up previous studies of the structure- function regulation of Hb oxygenation by anions Cl - and/or IHP. We measured the effect of these interactions on cooperativity and oxygen affinity, protons release upon oxygenation (Bohr Effect), and protein conformation. The role of specific residues on this effect was investigated trough results obtained with HbA, desArg Hemglobin and Hemoglobin Chesapeake (R92L), a natural mutant in which Arg- 92α is changed to Leu. The results showed that the suppression of C-terminal Arg-141 implies in non- cooperative O2-binding to Hemoglobin, lowering the protons released during oxygenation, and the protons release remains the same on chloride absence and presence. The removal of this residue didn´t affect the Bohr Effect on IHP presence. The results showed that O2 binding in the presence of IHP don´t induce quaternary structure transition observed in the absence of anions. Nevetheless, the cooperativity was maintained. The arg-141 have a main role on IHP stabilization of oxy and deoxy protein quaternary structure T. This conclusion is supported by results desArg Hemoglobin oxygenation determined in IHP presence, with and without NaCl. Calorimeters assays of IHP interaction with three Hemoglobin species, carried through Isothermal Titration Calorimetry confirm the Arg-141 importance on oxygenated... (Complete abstract click electronic access below)

Orientador: Marcio Francisco Colombo / Banca: Marcelo Matos Santoro / Banca: Flávio Augusto Vicente Seixas / Resumo: A ligação do O2 à Hemoglobina (Hb) é um processo regulado por interações alostéricas. Os íons Inositol hexafosfato (IHP) e cloreto, e prótons, são efetores alostéricos que estabilizam a Hb na estrutura quaternária T, diminuindo a afinidade da proteina pelo O2 em meio alcalino. A ligação de IHP também altera a cooperatividade de algumas hemoglobinas como, por exemplo a hemoglobina desArg, uma variante da HbAo da qual o resíduo arginina 141 das cadeia alfas foram retirados enzimaticamente. Estas argininas são prováveis sitios de interação com ions cloreto. Neste trabalho investigou-se a influência que a ligação de ânions Cl - e/ou IHP à Hb nas propriedades funcionais e conformacionais da proteína. Foi determinado o efeito destas interações na afinidade e cooperatividade de ligação de oxigênio à proteína, na liberação de prótons com a oxigenação ( efeito Bohr ), e na conformação da proteína. O papel de resíduos específicos nestes efeitos foi investigado comparando resultados obtidos com HbAo, Hb-desArg, e Hb Chesapeake (R92L), um mutante natural na qual a arginina 92 esta substituída por leucina. Os resultados mostraram que a retirada do resíduo C-terminal da cadeia alfa, arginina 141, suprimiu a cooperatividade da hemoglobina, diminui o número de prótons liberados durante a oxigenação, e que esta liberação de prótons foi a mesma na presença ou ausência de íons cloreto. Ainda, a retirada deste resíduo não interferiu no efeito Bohr dependente de IHP. Os resultados mostraram que a saturação de oxigênio ligados à HbA0 na presença de IHP não induziu a mudança na estrutura quaternária observada na ausência deste ânion, e na sua presença em meio com NaCl. Apesar disto, a cooperatividade da ligação é mantida. A arg-141α tem papel fundamental na estabilização pelo IHP da estrutura quaternária... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: The hemoglobin (Hb) oxygenation is a molecular process regulated by allosteric interactions. Inositol hexaphosphate (IHP), chloride and protons are heterotropic effectors that stabilizes the deoxy state, the quaternary structure T, decreasing O2 affinity in alkaline solution. IHP binding also modifies cooperativity of several Hbs, such as desArg Hemoglobin, a variant form of HbA which is obtained by enzymatic cleavage of the two Arg-141alfa residues. This residue has been identified to be a Cl - binding site on Hb. This work follows up previous studies of the structure- function regulation of Hb oxygenation by anions Cl - and/or IHP. We measured the effect of these interactions on cooperativity and oxygen affinity, protons release upon oxygenation (Bohr Effect), and protein conformation. The role of specific residues on this effect was investigated trough results obtained with HbA, desArg Hemglobin and Hemoglobin Chesapeake (R92L), a natural mutant in which Arg- 92α is changed to Leu. The results showed that the suppression of C-terminal Arg-141 implies in non- cooperative O2-binding to Hemoglobin, lowering the protons released during oxygenation, and the protons release remains the same on chloride absence and presence. The removal of this residue didn't affect the Bohr Effect on IHP presence. The results showed that O2 binding in the presence of IHP don't induce quaternary structure transition observed in the absence of anions. Nevetheless, the cooperativity was maintained. The arg-141 have a main role on IHP stabilization of oxy and deoxy protein quaternary structure T. This conclusion is supported by results desArg Hemoglobin oxygenation determined in IHP presence, with and without NaCl. Calorimeters assays of IHP interaction with three Hemoglobin species, carried through Isothermal Titration Calorimetry confirm the Arg-141 importance on oxygenated... (Complete abstract click electronic access below) / Mestre

As long ago as 1881, it was realized that a functional group of atoms in a molecule would cause an absorption band to appear at a particular frequency in the infrared spectrum of the molecule. In more recent years, the concept of characteristic group frequencies has become firmly established and has resulted in the present widespread use of infrared spectroscopy. There appear to have been relatively few studies of infrared absorption of organic acids as compared with their salts.

The problem with which this research is concerned is the determination of inorganic anions and cations with single injection ion chromatography. Direct detection of the separated analyte ions occurs after the analyte ions have passed through ion-exchange resins where they are separated according to their affinity for the ion-exchange resin active sites. The techniques involve the use of essentially a non-suppressed ion chromatographic system followed by a suppressed ion chromatographic system. With this system it is possible to accomplish both qualitative and quantitative determinations.

No description available.

The role that aquatic actinomycetes play, in the production of tastes and odors in water supplies has been investigated since 1948. The ability of these organisms to produce by-products in lakes and streams, which renders the water unpalatable, is of considerable public health importance. It is desirable that the waterworks industry has as much information as possible concerning the factors that contribute to the growth of these organisms. Since it appears that the aquatic actinomycetes may be isolated from most fresh-water sources, the problem of diversified environments and nutritional requirements offers an excellent field of investigation. The fresh-waters of the world contain variable quantities of electrolytes that may determine in part the biological activity of these organisms. The unsolved questions in this instance are concerned with the electrolytes present and their quantitative effects on the growth and development of these forms.