Navegando por Palavras-chave "Triton X-100"

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    Membrane permeabilization induced by Triton X-100: The role of membrane phase state and edge tension
    (Elsevier Ireland Ltd, 2017) Mattei, Bruno [UNIFESP]; Lira, Rafael Bezerra de [UNIFESP]; Perez, Katia Regina [UNIFESP]; Riske, Karin do Amaral [UNIFESP]
    Detergents are widely used to solubilize and separate biomembrane components. It is therefore relevant to study and understand the mechanistic details underlying detergent-lipid interactions using biomimetic systems. Here, we have investigated in detail the process of membrane permeabilization and the nature of pores induced by sub-solubilizing concentrations of the detergent Triton X-100 (TX-100) in bilayers composed of palmitoyl oleoyl phosphatidylcholine (POPC), sphingomyelin (SM) and binary mixtures of these phospholipids with 30 mol% cholesterol (chol). A fluorescence quenching assay was used to evaluate the permeability of large unilamellar vesicles (LUVs) in the presence of increasing concentrations of TX-100. Confocal microscopy was employed to visualize and quantify the permeability of giant unilamellar vesicles (GUVs) to two fluorescent dyes of different sizes in the presence of TX-100. Both methods showed that POPC, POPC/chol and SM membranes become fully permeable at a specific TX-100 concentration, followed by complete (POPC and SM) and partial (POPC/chol) solubilization at a higher detergent concentration. The confocal microscopy experiments revealed that opening of pores occurs as a well-defined event and that for POPC and POPC/chol the pores were initially selective to the small probe and then grew and allowed passage of the larger dye as well. On the other hand, the insoluble SM/chol membranes exhibited only a mild TX-100-induced permeabilization. The membrane edge tension of the liquid phases was measured from the closure rate of macropores induced by electric pulses in GUVs. Membrane edge tension was shown to be sensitive to membrane composition and to decrease in the presence of TX-100. We propose that extensive permeabilization occurs below a critical membrane edge tension, which is eventually reached in the partially and fully soluble compositions, but not in the insoluble mixture. (C) 2016 Elsevier Ireland Ltd. All rights reserved.
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    Morphological and physiological changes in Tetrahymena pyriformis for the in vitro cytotoxicity assessment of Triton X-100
    (Elsevier B.V., 2003-06-01) Dias, N.; Mortara, R. A.; Lima, N.; Univ Minho; Universidade Federal de São Paulo (UNIFESP)
    Non-ionic surfactants such as Triton X-100 have been widely used in industrial processing and in cleaning products for almost 50 years, being effective and economic emulsifying, wetting agents, dispersants and solubilizers. Cleaning products containing these surfactants are disposed of mainly by discharge into wastewater, which receives biological treatment in wastewater treatment systems. However, surface-active agents interact with eukaryotic cell membranes leading to biological damage at high concentrations. Tetrahymena pyriformis was used here as model organism to assess the effects of Triton X-100 through a series of in vitro cytotoxicity tests. Growth rates and morphological changes were, by their simplicity and reproducibility, the simplest toxicological assays. Cytoskeleton analysis seemed to be related with phagocytosis rate. Viability was evaluated by two different tests. Calcein AM/EthD-1 was used to assess T. pyriformis membrane damage during the 48-h experiment. the colorimetric MTT assay proved to be highly sensitive even at very short periods of Triton X-100 exposure. Tests performed in this study included simple and fast bioassays that provide overall information on the morphological and physiological state of cells exposed to different non-lytic and lytic concentrations of Triton X-100. (C) 2003 Elsevier B.V. All rights reserved.
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    Solubilização de membranas modelo de diferentes composições pelo detergente triton X-100
    (Universidade Federal de São Paulo (UNIFESP), 2017-03-29) Mattei, Bruno [UNIFESP]; Riske, Karin do Amaral [UNIFESP]; http://lattes.cnpq.br/9178927522709552; http://lattes.cnpq.br/4260343698344529; Universidade Federal de São Paulo (UNIFESP)
    Detergents are amphiphilic molecules widely used as solubilizing agents Biological membranes. However, biological membranes treated with detergents Ac-ese insoluble fragments rich in sphingolipids and cholesterol. Due to similarities of These features have been associated with lipid rafts and the liquid phase The. Here the study of the physical-chemical aspects that govern the process of UUilllCATiOn of membrane mimetic models: lipid vesicles of controlled composition Pure or mixtures, mainly POPC, SM and cholesterol). Many different Techniques were used to study the solubilization process of lipid vesicles by Detergent TX-100, one of the most used detergents in Biological membranes. The general phenomenon and the size of the structures formed during the Solubilization was obtained by turbidity, static and / or dynamic light scattering; The aspects Thermodynamic properties of the detergent-lipid interaction were investigated by titration calorimetry And the possible interference of the detergent in the phase transition of lipids was investigated Differential scanning calorimetry; The ability of detergents to permeabilize the The membrane was investigated by kinetics of leakage of a fluorescent probe and microscopy Confocal; The process of solubilization of giant vesicles was evaluated by optical microscopy of Phase contrast and fluorescence. In the work, different lipid compositions were investigated (Pure phospholipids, binary lipid mixtures and ternary lipid mixtures) in order to obtain Information on the factors that influence the obtaining of insoluble membrane fragments. Through the experiments it was possible to conclude that pure cell membranes (POPC and SM), reaching the liquid-disordered phase and gel are totally soluble to the action of Detergent, whereas the addition of cholesterol, inducing a higher degree of Lipids leads to conditions in which the membranes are partially or totally insoluble; The factor of Insolubility is conditioned to cholesterol concentration in vesicles with binary mixtures Of lipids, reaching total insolubility in situations in which the membrane tends or reaches the Liquid-ordered phase; This is also reflected in the detergent affinity constant for these Membranes. In addition, vesicles in the gel phase are solubilized with lower concentrations of TX-100 than vesicles in the liquid-disordered phase, possibly due to the lower Ability of the membrane to accommodate detergents; Added to this, the addition of detergent Also appears to induce a phase transition in vesicles in the gel phase and a coexistence of Fluid phases in POPC: cholesterol vesicles. In ternary lipid mixtures, the situations in Higher solubility resistance is observed in regions with Greater fraction of SM and cotesterol. Finally, the membranes become permeable in TX-100 / lipid molars lower than the solubilization process, showing that even before To reach the bi-co-existent phase of the membrane, the detergent already changes the membrane structure.