Origem da estabilização de eritrócitos por sorbitol
| Ano de defesa: | 2006 |
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| Autor(a) principal: | |
| Orientador(a): | |
| Banca de defesa: | |
| Tipo de documento: | Dissertação |
| Tipo de acesso: | Acesso aberto |
| Idioma: | por |
| Instituição de defesa: |
Universidade Federal de Uberlândia
BR Programa de Pós-graduação em Genética e Bioquímica Ciências Biológicas UFU |
| Programa de Pós-Graduação: |
Não Informado pela instituição
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| Departamento: |
Não Informado pela instituição
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| País: |
Não Informado pela instituição
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| Palavras-chave em Português: | |
| Link de acesso: | https://repositorio.ufu.br/handle/123456789/15835 |
Resumo: | ABSTRACT - CHAPTER I With the aim of guarantee the stability of the biological organization complexes, nature uses several mechanisms, as the control of pH, temperature and concentration of solutes in the internal medium. The control of the solute concentration contributes to what we can call as osmostabilization. This paper tries to apply the known theories about the stabilization of proteins now on the biological membranes. The osmostabilizing solutes, also called as osmolytes, increase the free energy of the native state (N), but increase much more the free energy of the unfolded state (D) of a protein, in such a way that the osmolytes stabilize the N state of a protein by the increase in free energy barrier between the states, which makes less probable the unfolding and the loss of the protein function. The reason of this difference would be in the preferential interaction of the protein with the water. The protein prefers binds to water than to the osmolyte, which is then excluded from the inner hydration shell of the protein surface, according an effect that was designated as solvophobic or osmophobic effect. The larger contact surface of the D state requires a higher investment of free energy for its hydration, beyond a better organization of the water molecules around the external surface, in such a way that the conformational entropy of the solvent is lower around the D state than it is around the N state. This means that the osmolytes stabilize the native state in relation to the unfolded state of the protein. The biological membranes have an important aspect of their structures in common with water soluble proteins. They also hide their hydrophobic groups in their anhydrous interior, constituted by a phospholipids bilayer, in which the polar heads of these lipids make contact with the aqueous surrounding in the external and internal media of the biological structure encompassed by the membrane. The erythrocytes constitute a very valid model to study the behavior of the membranes. The utilization of osmolytes increases the stability of erythrocytes preparations and permits even their cryopreservation. In the presence of osmolytes the erythrocytes suffer reversible morphological alterations with volume decrease. Thus, the erythrocytes will exist in an equilibrium process between two states, an expanded (R) and a compact one (T), where T is the more stable state. In this study, we tentatively explained this larger stability of the T state of the erythrocytes with basis in the preferential hydration theory of Timasheff. ABSTRACT - CHAPTER II The erythrocyte constitutes a very adequate model to study the stability of biological membranes, since the rupture of its membrane promotes release of hemoglobin, which capacity to adsorb light in the visible region of the spectra permits the monitoration of the membrane denaturation. In this work, we studied the effect of the presence of sorbitol on the thermal dependence of the stability of human erythrocytes against the denaturant action of ethanol in 0,9% NaCl. The membrane denaturation was monitored by the measurement of the absorbance at 540 nm (A540 nm). The dependence of A540 nm with the ethanol concentration in 0.9% NaCl, in the absence and presence of 1 mol.L-1 sorbitol, was studied at 27, 32, 37 and 42 °C. After complete denaturation of the erythrocytes, the A540 nm values were converted in percentage of hemolysis. All the dependencies of the % of hemolysis with the concentration of ethanol followed sigmoidal transition lines, which were adjusted to the Boltzman equation, in order to determine the concentration of ethanol able to promote 50% of hemolysis (D50). The incorporation of 1 mol.L-1 sorbitol promoted statistically significant decreases (P<0.01) in the D50 values, for all considered temperatures. The increase in the temperature also promoted statistically significant decreases (P<0.01) in the D50 values in the absence and in the presence of sorbitol. The values of D50 presented a linear dependence with the temperature. The slope of that line in the presence of sorbitol was significantly smaller (P<0.01) than it was in absence of that solute. This means that the presence of 1 mol.L-1 sorbitol increases the chaotropic action of ethanol, at the same time that it presents a stabilizing action, which increase with an increase in the temperature. If we assume linearity beyond the interval of 27 and 42 °C, those regression lines will intercept around 68.8 °C, where the stabilizing effect of sorbitol would neutralize its synergism with the chaotropic action of ethanol. These effects were explained with basis in a two state equilibrium model for the erythrocyte, a less stable expanded state and a more stable contracted state. The rationality of the model is discussed. Whatever is the adopted explanation, our results permit conclude that 1 mol.L-1 sorbitol, in the presence of 0.9% NaCl, increases the chaotropic action of ethanol and temperature on the erythrocyte membrane, although it does not present any chaotropic action itself between 0 and 1.5 mol.L-1, by the same time that it also presents a stabilizing action on the membrane that increases with the increase in the temperature. |