Novel Two-Band Ratiometric Fluorescence Probes with Different Location and Orientation in Phospholipid Membranes (original) (raw)
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FEBS Letters, 2001
We report on dramatic differences in fluorescence spectra of 4P P-dimethylamino-3-hydroxyflavone (probe F) studied in phospholipid membranes of different charge (phosphatidyl glycerol, phosphatidylcholine (PC), their mixture and the mixture of PC with a cationic lipid). The effect consists in variations of relative intensities at two well-separated band maxima at 520 and 570 nm belonging to normal (N*) and tautomer (T*) excited states of flavone chromophore. Based on these studies we propose a new approach to measure electrostatic potential at the surface layer of phospholipid membranes, which is based on potential-dependent changes of bilayer hydration and involves very sensitive and convenient ratiometric measurements in fluorescence emission. ß 2001 Published by Elsevier Science B.V. on behalf of the Federation of European Biochemical Societies.
Excited state proton transfer fluorescence of 3-hydroxyflavone in model membranes
Spectrochimica Acta Part A-molecular and Biomolecular Spectroscopy, 1997
3-Hydroxyflavone (3HF), the basic structural moiety of an important group of naturally occurring, biologically active flavonoids, has attracted extensive attention for its intramolecular excited-state proton-transfer (ESPT) and dual fluorescence characteristics. We report here, for the first time, its fluorescence (emission, excitation, polarization anisotropy and lifetime) behaviour in model membranes consisting of small, unilamellar liposomes of synthetic phosphatidylcholine (dimyristoyl phosphatidylcholine (DMPC) and dipalmitoyl phosphatidylcholine (DPPC)). The emission spectrum is conspicuously dominated by the ESPT tautomer fluorescence band. This result, along with other relevant data, indicates that the molecules are embedded in the lipid bilayer matrix facing environments where external H-bonding perturbation effects are minimized. The anisotropy (Y) versus temperature (r) profiles dramatically reveal the thermotropic gel-liquid crystalline phase transition properties of the phospholipids. signifying a novel application of the intrinsic (tautomer) fluorescence of 3HF. 0 1997 Elsevier Science B.V.
2014
Indian Chemical Society, 92, Acharya Prafulla Chandra Road, Kolkata-700 009, India <em>E-mail</em> : drrahulchem@rediffmail.com Department of Chemistry, West Bengal State University (Barasat), Kolkata-700 126, India <em>E-mail </em>: ranjan.das68@gmail.com <em>Manuscript received online 07 December 2012, revised 09 January 2013, accepted 11 March 2013</em> <strong>The photophysics of a dye, <em>N</em>-[[4'-<em>N,N</em>-diethylamino-3-hydroxy-6-flavonyl]methyl]-<em>N</em>-methyl-<em>N</em>-(3-sulfopropyl)- 1-dodecanaminium, inner salt (F2N12S), in the large unilamellar vesicles of neutral egg yolk phosphatidylcholine (EYPC) and negatively charged egg yolk phosphatidylglycerol (EYPG) lipids were investigated by steady state and time-resolved fluorescence spectroscopy. The fluorescence spectra display a dramatic variation of the relative intensity of dual emission bands attributed to the norma...
Bimodal Distribution and Fluorescence Response of Environment-Sensitive Probes in Lipid Bilayers
Biophysical Journal, 2004
A remarkable heterogeneity is often observed in the spectroscopic properties of environment-sensitive fluorescence probes in phospholipid bilayers. To explain its origin, we provided a detailed investigation of the fluorescence excitation and emission spectra of 4#-dimethylamino-3-hydroxyflavone (probe F) in bilayer vesicles with the variations of fatty acid composition, polar heads, temperature, and cholesterol content. Probe F, due to excited-state intramolecular proton transfer, exhibits two bands in emission that are differently sensitive to intermolecular interactions-thereby allowing us to distinguish universal (dipole-dipole) and specific (H-bonding) interactions within the bilayer. Spectroscopic, quenching, and anisotropy data suggest the presence of two forms of probe F at different locations in the bilayer: an H-bond free form located below sn 1 -carbonyls and an H-bonded form located at the polar membrane interface. We provide a quantitative analysis of the distribution of the probe between these two locations as well as the polarity of these locations, and show that both the distribution and the polarity contribute to the probe response. Moreover, analysis of literature data on other environment-sensitive probes (Prodan, Laurdan, Nile Red, NBD lipids, etc.) in lipid bilayers allows us to suggest that the bimodal distribution in the lipid bilayer is probably a general feature of low-polar molecules with polar groups capable of H-bonding interactions.
Cell Biochemistry and Biophysics, 2013
Two novel precursors of fluorescent dyes (PFD813 and PFD814) have been studied for their ability to photo-activation, transfer across the biomembrane and cells staining. The fluorescent dyes Rho813 and Rho814 formed by photo-activation of their precursors PFD813 and PFD814 inside cells were used for the optical detection of particular features in vitro (HaCat cells, human epithelial carcinoma A431, epidermoid carcinoma of the cervix HeLa and chinese hamster ovary CHO cells). One of the possibilities to visualize and track the pathways of macromolecules or organelles in a "living" cell is to monitor them after staining with these PFDs during the real time measurements. A bright fluorescent signal from the photoactivated dye molecules inside the small spot in the cell can be monitored during their movement into the cell dark region (where the dye was not activated and did not fluoresce). The obtained data are important for further application of these precursors of the fluorescent dyes ("caged" dyes) for microscopic probing of biological objects.
Biochimica et Biophysica Acta (BBA) - Biomembranes, 2004
The penetration of water into the hydrophobic interior leads to polarity and hydration profiles across lipid membranes which are fundamental in the maintenance of membrane architecture as well as in transport and insertion processes into the membrane. The present paper is an original attempt to evaluate simultaneously polarity and hydration properties of lipid bilayers by a fluorescence approach. We applied two 3-hydroxyflavone probes anchored in lipid bilayers at a relatively precise depth through their attached ammonium groups. They are present in two forms: either in H-bond-free form displaying a two-band emission due to an excited state intramolecular proton transfer reaction (ESIPT), or in H-bonded form displaying a single-band emission with no ESIPT. The individual emission profiles of these forms were obtained by deconvolution of the probes' fluorescence spectra. The polarity of the probe surrounding the bilayer was estimated from the two-band spectra of the H-bond-free form, while the local hydration was estimated from the relative contribution of the two forms. Our results confirm that by increasing the lipid order (phase transition from fluid to gel phase, addition of cholesterol or decrease in the lipid unsaturation), the polarity and to a lesser extent, the hydration of the bilayers decrease simultaneously. In contrast, when fluidity (i.e. lipid order) is kept invariant, increase of temperature and of bilayer curvature leads to a higher bilayer hydration with no effect on the polarity. Furthermore, no correlation was found between dipole potential and the hydration of the bilayers.
Journal of Fluorescence, 2009
3-Hydroxyquinolones (3HQs) are a new class of water soluble dual fluorescence probes that can monitor both polarity and basicity (H-bond accepting ability) parameters. Both parameters play an important role in proteins and lipid membranes. Nevertheless, no method exists actually to measure the basicity parameter separately from the polarity. To achieve this aim, we synthesized 2benzofuryl-3-hydroxy-4(1H)-quinolone (3HQ-Bf) and characterized its photophysical properties by UV, steady-state and time-resolved fluorescence spectroscopy. Due to its extended conjugation and totally planar conformation, 3HQ-Bf is characterized by a high fluorescence quantum yield. In solution, this dye shows an excited state intramolecular proton transfer (ESIPT) reaction resulting in two tautomer bands in the emission spectra. The ESIPT reaction can be considered as irreversible and is governed by rate constants from 0.6 to 8×10 9 s −1 , depending on the solvent. The analysis of the spectral properties of 3HQ-Bf in a series of organic solvents revealed a marginal sensitivity to the solvent polarity, but an exquisite sensitivity to solvent basicity, as shown by the linear dependence of the logarithm of the emission bands intensity ratio, log(I N* /I T* ), as well as the absorption or emission maxima wavenumbers as a function of the solvent basicity parameter. This probe may find useful applications through coupling to a protein ligand, for characterizing the H-bond acceptor ability at the ligand binding site as well as for studying the basicity changes of lipid membranes during their chemo-and thermotropic conversions.
Journal of biochemical and biophysical methods, 2005
We report on the synthesis by coupling of a triterpenoid oleanolic acid with 4V-diethylamino-3-hydroxyflavone (FE) to produce an environment-sensitive biomembrane probe with two-band ratiometric response in fluorescence emission. The synthesized compound (probe FOT) was tested in a series of model solvents and demonstrated the response to solvent polarity and intermolecular hydrogen bonding very similar to that of parent probe FE. Meantime when incorporated into lipid bilayer membranes, it showed new features differing in response between lipids of different surface charges as well as between glycerophospholipids and sphingomyelin. 0165-022X/$ -see front matter D We observed that in the conditions of coexistence of rafts and non-raft structures the probe is excluded from the rafts. D