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|Title:||Quantification of Cholesterol Solubilization by Bile Salt Micelles: Implications in its Intestinal Absorption by Passive Processes||Authors:||Gomes, Filipe Coreta||Orientador:||Silvestre, Maria||Issue Date:||14-Jan-2014||Citation:||GOMES, Filipe Manuel Coreta - Quantification of cholesterol solubilization by bile salt micelles : implications in its intestinal absorption by passive processes. Coimbra : [s.n.], 2013. Tese de doutoramento. Disponível na WWW: http://hdl.handle.net/10316/23830||Abstract:||Cholesterol absorption takes place at the intestine. Prior to absorption, and due to its low aqueous solubility, cholesterol must be solubilized in bile salts (BS). Cholesterol is an important molecule for several functions from structural to hormonal. However, excess of cholesterol is implicated in several diseases, namely atherosclerosis and gallstone disease. The understanding of the mechanism beneath cholesterol absorption is relevant for the effectiveness of strategies to decrease cholesterol intake from the intestinal lumen. In this work, focus was given to understand some of the principles related to cholesterol solubilization by BS as well as the interaction of BS with model membranes. First we explored the cholesterol solubilization by glycodeoxycholic acid (GDCA), glycochenodeoxycholic acid (GCDCA) and glycocholic acid (GCA) in micelles of single, binary and ternary mixtures, as these are the most prevalent BS at intestinal lumen. Results showed that single micelles of the most hydrophobic BS (GDCA) presented the highest cholesterol solubilization, followed by micelles of GCDCA and of GCA. The 3 compositions of binary micelles, GCA/GCDCA (50:50), GCA/GDCA (50:50) and GCDCA/GDCA (50:50), and the ternary mixture composed of GCA/GCDCA/GDCA (40/40/20) presented lower solubilization than the simple micelles of GDCA but higher than GCDCA micelles. The results from this study suggest that BS mixtures present at duodenum are optimized neither for the highest nor for the lowest solubility of cholesterol; instead the solubilization capacity can be understood as a tradeoff between cholesterol supply to membranes and cholesterol excretion. The results showed a linear correlation between cholesterol solubilization by BS micelles and the total concentration of cholesterol, until a plateau be reached, indicating the cholesterol saturation index (CSI). Moreover, the sterol occupancy per micelle differs on the BS, being 1 to simple micelles of GDCA and inferior to the ones of GCDCA and GCA. The interpretation of these results leads to the understanding of solubilization of cholesterol by BS micelles as a partition phenomena rather than a binding event to an available place at the micelle. Solubilization is higher or lower depending on the partition of the molecule considered to the micelle. Once the solubility of this molecule is attained no further solubilization of this molecule will occur however, this does not mean that micelle doesn’t maintain solubility capacity for other molecules. The understanding of the solubilization phenomena by the micelles in the presence of several co-solutes can give important information on strategies to decrease cholesterol solubilization and absorption. Pursuing this objective, the effect of stigmasterol and stigmastanol (phytosterols), palmitic acid (saturated fatty acid), oleic acid (unsaturated fatty acid) and α-tocopherol (vitamin) on the solubilization of cholesterol by the BS micelles was addressed. The highest effect in the decrease of cholesterol solubility was observed in the presence of phytosterols, followed by the α-tocopherol and very small effect of oleic acid. Phytosterols were solubilized in BS micelles until their limit of solubilization was attainded, while α-tocopherol and oleic acid were soluble in the range of concentration used. For phytosterols, once reached their limit of solubilization at micelle, increasing their concentration resulted in a decrease of cholesterol solubilization in the case of stigmasterol, while no effect in the case of stigmastanol. This suggests that stigmastanol, α-tocopherol and oleic acid exhibit a co-solubilization mechanism, while stigmasterol exhibits both co-solubilization and co-precipitation. Unlikely, the presence of palmitic acid in the micelles as co-solute enhance cholesterol solubility. Looking at the absorption phenomena we addressed the partition of several BS to model membranes. From the results, it was clear that the most hydrophobic BS partitioned more to POPC and to PC/SpM/Chol (1:1:1) membranes than the most hydrophilic. Also partition to this latter membrane was lower that to POPC. Qualitative data was obtained for translocation in POPC membranes, showing higher values for the most hydrophilic BS, both in conjugated and non conjugated BS. The translocation of BS into POPC/SpM/Chol membranes showed to be faster for both BS measured (DCA and CDCA). The enthalpy of partition showed a negative value for the partition of BS to liquid disordered phase, like in the case of POPC, and showed a positive enthalpy of partition for the membranes of POPC/SpM/Chol. The establishment of the driving rules in the process of solubilization and partition to membranes can give an important clue on a better understanding of difusion process at the intestine.||Description:||Tese de doutoramento em Quimica, na especialidade de Quimica Biológica, apresentada à Faculdade de Ciências e Tecnologia da Universidade de Coimbra||URI:||http://hdl.handle.net/10316/23830||Rights:||openAccess|
|Appears in Collections:||FCTUC Química - Teses de Doutoramento|
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