Sonia Freire - Academia.edu (original) (raw)

Papers by Sonia Freire

Dyes and Pigments, 2014

Thioflavin T is a highly sensitive fluorescent marker of amyloid fibrils that has been widely use... more Thioflavin T is a highly sensitive fluorescent marker of amyloid fibrils that has been widely used for in vitro biomedical assays. However, neither its complex photophysical behavior nor its binding mode to amyloid fibrils are still well understood. We present a detailed analysis of the photophysical properties of Thioflavin T in various media, including solvents and solvent mixtures of different viscosities as well as fibrillar and globular proteins. We propose a model that explains the strong wavelength dependency of the Thioflavin T fluorescence and the large fluorescence enhancement in certain environments. We determine the binding affinities and the fluorescence properties of Thioflavin T bound to amyloid-b (1 e42) fibrils and to bovine serum albumin and discuss the sensitivity and the specificity of this probe to amyloid aggregates. These results allow us to assess the suitability of Thioflavin T for quantitative determinations in biomedical studies.

Chemistry - A European Journal, 2015

The aggregation of amyloid-β peptide and its accumulation in the human brain has an important rol... more The aggregation of amyloid-β peptide and its accumulation in the human brain has an important role in the etiology of Alzheimer's disease. Thioflavin T has been widely used as a fluorescent marker for these amyloid aggregates. Nevertheless, its complex photophysical behavior, with strong wavelength dependencies of all its fluorescence properties, requires searching for new fluorescent probes. The use of 2-(2'-hydroxyphenyl)imidazo[4,5-b]pyridine (HPIP), which shows two emission bands and a rich excited-state behavior due to the existence of excited-state intramolecular processes of proton transfer and charge transfer, is proposed. These properties result in a high sensitivity of HPIP fluorescence to its microenvironment and cause a large differential fluorescence enhancement of the two bands upon binding to aggregates of the amyloid-β peptide. Based on this behavior, a very sensitive ratiometric method is established for the detection and quantification of amyloid fibrils, which can be combined with the monitoring of fluorescence anisotropy. The binding selectivity of HPIP is discussed on the basis of the apparent binding equilibrium constants of this probe to amyloid-β (1-42) fibrils and to the nonfibrillar protein bovine serum albumin. Finally, an exhaustive comparison between HPIP and thioflavin T is presented to discuss the sensitivity and specificity of these probes to amyloid aggregates and the significant advantages of the HPIP dye for quantitative determinations.

Soft Matter, 2013

A quantitative empirical model is presented, which relates the total surfactant concentration wit... more A quantitative empirical model is presented, which relates the total surfactant concentration with the fluorescence intensity and the mean translational diffusion coefficient of dyes in micellar solutions, as determined by Fluorescence Correlation Spectroscopy (FCS). Based on this model a systematic data analysis method is defined for the precise determination of the dye binding equilibrium constant, the critical micelle concentration, the translational diffusion coefficients, and the hydrodynamic radii of dyes 10 and micelles and related properties. The method can be used for the routine and automatic determination of the cmc of surfactant solutions. The model is applied to dyes with very different hydrophobicity in aqueous solutions of the non-ionic surfactant Triton X-100. As a reference the cmc of Triton X-100 is also determined directly from changes in the absorbance of its phenyl-ring applying a ratiometric method. 65 characterization of a surfactant solution, for example for the determination of the cmc, it is imperative to take the dye exchange equilibrium into account.

Photochemical & Photobiological Sciences, 2010

The nature and strength of the interactions between a cationic fluorophore, Rhodamine 123 (R123),... more The nature and strength of the interactions between a cationic fluorophore, Rhodamine 123 (R123), and surfactants of different head charge are investigated. Series of absorption and fluorescence emission spectra and of fluorescence decays are measured. R123 does not interact with the monomers of both nonionic and cationic surfactants but it presents affinity to their micelles. A partition equilibrium model was proposed and the corresponding equilibrium constants were obtained, as well as the photophysical properties of the dye bound to the micelles. In the case of the cationic surfactants, changes of the fluorescence properties were already observed below the critical micellar concentration (CMC) due to dynamic quenching caused by the free counterions. In the presence of anionic surfactants R123 shows a very different behaviour with dramatic spectral changes below the CMC. The observed variations are attributed to a first, strong interaction between R123 and the surfactant monomers, which yields ionic pairs of singular photophysical properties and dominates at low surfactant concentrations, followed by the association of R123 with the surfactant premicellar and micellar aggregates at higher surfactant concentrations near the CMC. This behaviour results from the competition between the strong electrostatic interactions of the cationic dye with the anionic surfactant head groups and the hydrophobic forces stabilizing the dye inside the micelles. The results of this work illustrate the complex physicochemical and photophysical behaviour of a charged dye in micellar systems, which resembles the expected situation in similar systems such as biological membranes.

We have studied the complexation between an adamantane derivative labelled with the fluorescent p... more We have studied the complexation between an adamantane derivative labelled with the fluorescent probe Alexa 488 and the three natural cyclodextrins (-, and -CD) by Fluorescence Correlation Spectroscopy (FCS), demonstrating the ability of this technique to detect association and to determine the corresponding equilibrium constants with no need for changes in the fluorescence properties of the guest. At low CD concentrations the observed increase of the diffusion time of the probe is mainly due to the complexation of the adamantyl moiety, but further changes are observed when increasing CD concentration that are attributed to the formation of CD aggregates. These aggregates appear at quite low CD concentrations and seem to be formed by a small number of CD molecules. These results show the potential of FCS for the study of CD self-assembly, a recently-recognized phenomenon that could be used to improve certain applications of CDs.

The role of the host size on stoichiometry, structure and strength of supramolecular association ... more The role of the host size on stoichiometry, structure and strength of supramolecular association between a fluorescent probe, coumarin 460 (C460), and the three natural cyclodextrins (CDs) with increasing cavity sizes (α, β fluorescent probe, coumarin 460 (C460), and the three natural cyclodextrins (C s) with increasing cavity si es (α, β and γ-cyclodextrin) is analyzed, using fluorescence spectroscopy. The variations of the fluorescence emission spectra of C460 with CD concentration can be explained with a 1:1 complexation equilibrium model for α-CD and β-CD, but also a 1:2 complex has to be included in the case of γ-CD. Nevertheless, the photophysical properties and the stability of the complexes do not correlate in a simple way with the cavity size. For instance, the association between y p p y y , C460 and β-CD, which is the CD with medium cavity size, is much stronger than with the other CDs but it does not provoke fluorescence enhancement as in those cases. Detailed analysis o...

Dyes and Pigments, 2014

Thioflavin T is a highly sensitive fluorescent marker of amyloid fibrils that has been widely use... more Thioflavin T is a highly sensitive fluorescent marker of amyloid fibrils that has been widely used for in vitro biomedical assays. However, neither its complex photophysical behavior nor its binding mode to amyloid fibrils are still well understood. We present a detailed analysis of the photophysical properties of Thioflavin T in various media, including solvents and solvent mixtures of different viscosities as well as fibrillar and globular proteins. We propose a model that explains the strong wavelength dependency of the Thioflavin T fluorescence and the large fluorescence enhancement in certain environments. We determine the binding affinities and the fluorescence properties of Thioflavin T bound to amyloid-b (1 e42) fibrils and to bovine serum albumin and discuss the sensitivity and the specificity of this probe to amyloid aggregates. These results allow us to assess the suitability of Thioflavin T for quantitative determinations in biomedical studies.

Chemistry - A European Journal, 2015

The aggregation of amyloid-β peptide and its accumulation in the human brain has an important rol... more The aggregation of amyloid-β peptide and its accumulation in the human brain has an important role in the etiology of Alzheimer's disease. Thioflavin T has been widely used as a fluorescent marker for these amyloid aggregates. Nevertheless, its complex photophysical behavior, with strong wavelength dependencies of all its fluorescence properties, requires searching for new fluorescent probes. The use of 2-(2'-hydroxyphenyl)imidazo[4,5-b]pyridine (HPIP), which shows two emission bands and a rich excited-state behavior due to the existence of excited-state intramolecular processes of proton transfer and charge transfer, is proposed. These properties result in a high sensitivity of HPIP fluorescence to its microenvironment and cause a large differential fluorescence enhancement of the two bands upon binding to aggregates of the amyloid-β peptide. Based on this behavior, a very sensitive ratiometric method is established for the detection and quantification of amyloid fibrils, which can be combined with the monitoring of fluorescence anisotropy. The binding selectivity of HPIP is discussed on the basis of the apparent binding equilibrium constants of this probe to amyloid-β (1-42) fibrils and to the nonfibrillar protein bovine serum albumin. Finally, an exhaustive comparison between HPIP and thioflavin T is presented to discuss the sensitivity and specificity of these probes to amyloid aggregates and the significant advantages of the HPIP dye for quantitative determinations.

Soft Matter, 2013

A quantitative empirical model is presented, which relates the total surfactant concentration wit... more A quantitative empirical model is presented, which relates the total surfactant concentration with the fluorescence intensity and the mean translational diffusion coefficient of dyes in micellar solutions, as determined by Fluorescence Correlation Spectroscopy (FCS). Based on this model a systematic data analysis method is defined for the precise determination of the dye binding equilibrium constant, the critical micelle concentration, the translational diffusion coefficients, and the hydrodynamic radii of dyes 10 and micelles and related properties. The method can be used for the routine and automatic determination of the cmc of surfactant solutions. The model is applied to dyes with very different hydrophobicity in aqueous solutions of the non-ionic surfactant Triton X-100. As a reference the cmc of Triton X-100 is also determined directly from changes in the absorbance of its phenyl-ring applying a ratiometric method. 65 characterization of a surfactant solution, for example for the determination of the cmc, it is imperative to take the dye exchange equilibrium into account.

Photochemical & Photobiological Sciences, 2010

The nature and strength of the interactions between a cationic fluorophore, Rhodamine 123 (R123),... more The nature and strength of the interactions between a cationic fluorophore, Rhodamine 123 (R123), and surfactants of different head charge are investigated. Series of absorption and fluorescence emission spectra and of fluorescence decays are measured. R123 does not interact with the monomers of both nonionic and cationic surfactants but it presents affinity to their micelles. A partition equilibrium model was proposed and the corresponding equilibrium constants were obtained, as well as the photophysical properties of the dye bound to the micelles. In the case of the cationic surfactants, changes of the fluorescence properties were already observed below the critical micellar concentration (CMC) due to dynamic quenching caused by the free counterions. In the presence of anionic surfactants R123 shows a very different behaviour with dramatic spectral changes below the CMC. The observed variations are attributed to a first, strong interaction between R123 and the surfactant monomers, which yields ionic pairs of singular photophysical properties and dominates at low surfactant concentrations, followed by the association of R123 with the surfactant premicellar and micellar aggregates at higher surfactant concentrations near the CMC. This behaviour results from the competition between the strong electrostatic interactions of the cationic dye with the anionic surfactant head groups and the hydrophobic forces stabilizing the dye inside the micelles. The results of this work illustrate the complex physicochemical and photophysical behaviour of a charged dye in micellar systems, which resembles the expected situation in similar systems such as biological membranes.

We have studied the complexation between an adamantane derivative labelled with the fluorescent p... more We have studied the complexation between an adamantane derivative labelled with the fluorescent probe Alexa 488 and the three natural cyclodextrins (-, and -CD) by Fluorescence Correlation Spectroscopy (FCS), demonstrating the ability of this technique to detect association and to determine the corresponding equilibrium constants with no need for changes in the fluorescence properties of the guest. At low CD concentrations the observed increase of the diffusion time of the probe is mainly due to the complexation of the adamantyl moiety, but further changes are observed when increasing CD concentration that are attributed to the formation of CD aggregates. These aggregates appear at quite low CD concentrations and seem to be formed by a small number of CD molecules. These results show the potential of FCS for the study of CD self-assembly, a recently-recognized phenomenon that could be used to improve certain applications of CDs.

The role of the host size on stoichiometry, structure and strength of supramolecular association ... more The role of the host size on stoichiometry, structure and strength of supramolecular association between a fluorescent probe, coumarin 460 (C460), and the three natural cyclodextrins (CDs) with increasing cavity sizes (α, β fluorescent probe, coumarin 460 (C460), and the three natural cyclodextrins (C s) with increasing cavity si es (α, β and γ-cyclodextrin) is analyzed, using fluorescence spectroscopy. The variations of the fluorescence emission spectra of C460 with CD concentration can be explained with a 1:1 complexation equilibrium model for α-CD and β-CD, but also a 1:2 complex has to be included in the case of γ-CD. Nevertheless, the photophysical properties and the stability of the complexes do not correlate in a simple way with the cavity size. For instance, the association between y p p y y , C460 and β-CD, which is the CD with medium cavity size, is much stronger than with the other CDs but it does not provoke fluorescence enhancement as in those cases. Detailed analysis o...