Piotr Bojarski

Different ways to insert carotenoids into liposomes affect structure and dynamics of the bilayer differently.

We apply and quantify two techniques to incorporate carotenoids into liposomes: (i). preparation of unilamellar liposomes from mixtures of phospholipids and a carotenoid or cholesterol; (ii). insertion of carotenoids into prepared liposomes. Homogeneous liposomal fractions with a vesicle size diameter of approximately 50 nm were obtained by an extrusion method. The resulting vesicles were subjected to a three-dimensional light scattering cross-correlation measurement in order to evaluate their size distribution. The fluorescent dyes Laurdan, DiI-C(18), C(6)-NBD-PC were used to label the liposomes and to evaluate modulations of ordering, hydrophobicity and permeability to water molecules adjacent to the bilayer in the presence of carotenoids and/or cholesterol. Zeaxanthin incorporation (up to 0.1-1 mol%) attributes to the symmetric and ordered structure of the bilayer, causing both a strong hydrophobicity and a lower water permeability at the polar region of the membrane. The incorporation of lutein has similar effects, but its ordering effect is inferior in the polar region and superior in the non-polar region of the membrane. beta-Carotene, which can be incorporated at lower effective concentrations only, distributes in a more disordered way in the membrane, but locates preferentially in the non-polar region and, compared to lutein and zeaxanthin, it induces a less ordered structure, a higher hydrophobicity and a lower water permeability on the bilayer.

Tamoxifen perturbs lipid bilayer order and permeability: comparison of DSC, fluorescence anisotropy, Laurdan generalized polarization and carboxyfluorescein leakage studies

The perturbation of the lipid bilayer structure by tamoxifen may contribute to its multiple mechanisms of anticancer action not related to estrogen receptors. This study evaluates the effect of tamoxifen on structural characteristics of model membranes using differential scanning calorimetry (DSC), fluorescence anisotropy of 1,6-diphenyl-1,3,5-hexatriene (DPH) and 1-[4-[trimethylammonium)phenyl]-6-phenylhexa-1,3,5-triene (TMA-DPH), as well as 6-dodecanoyl-2-dimethylaminonaphthalene (Laurdan) generalized polarization. The comparative measurements in multilammelar vesicles (MLV) prepared from dipalmitoylphosphatidylcholine (DPPC) revealed that tamoxifen decreases the phase transition temperature (Tm) paralleled by a broadening of the phase transition profile. In large unilamellar vesicles (LUV) prepared from egg yolk phosphatidylcholine (EPC), tamoxifen increased the lipid bilayer order predominantly in the outer bilayer region. From membrane permeability measurements, we conclude that the tamoxifen-induced release of entrapped carboxyfluorescein (CF) results from a permanent bilayer disruption and the formation of transient holes in the lipid bilayer.

Concentration quenching and depolarization of rhodamine 6G in the presence of fluorescent dimers in polyvinyl alcohol films

Abstract Nonradiative energy transport in a concentrated system of rhodamine 6G in polyvinyl alcohol films is studied. Concentration changes in absorption and fluorescence spectra evidence the presence of rhodamine 6G fluorescent dimers. The results of concentration quenching and depolarization agree with the hopping theory of forward energy transport up to intermediate concentrations. At the highest concentrations, the experimental quantum yield is higher than the theoretical one and no repolarization effect is found at room temperature. These discrepancies are explained by the formation of fluorescent dimers which are imperfect traps for excitation energy. This conclusion is supported by measurements at elevated temperature.

Nonradiative excitation energy transport in one-component disordered systems.

AbstractHigh-accuracy Monte Carlo simulations of the time-dependent excitation probabilityGs(t) and steady-state emission anisotropyrM/r0M for one-component three-dimensional systems were performed. It was found that the values ofrM/r0M obtained for the averaged orientation factor

Long-distance FRET analysis: A Monte Carlo simulation study

\overline {\kappa ^2 }

Excitation energy transport and trapping in concentrated solid solutions of flavomononucleotide

only slightly overrate those obtained for the real values of the orientation factor κik2. This result is essentially different from that previously reported. Simulation results were compared with the probability coursesGs(t) andR(t) obtained within the frameworks of diagrammatic and two-particle Huber models, respectively. The results turned out to be in good agreement withR(t) but deviated visibly fromGs(t) at long times and/or high concentrations. Emission anisotropy measurements on glycerolic solutions of Na-fluorescein and rhodamine 6G were carried out at different excitation wavelengths. Very good agreement between the experimental data and the theory was found, with λex≈λ0-0 for concentrations not exceeding 3.5·10−2 and 7.5·10−3M in the case of Na-fluorescein and rhodamine 6G, respectively. Up to these concentrations, the solutions investigated can be treated as one-component systems. The discrepancies observed at higher concentrations are caused by the presence of dimers. It was found that forλex λ0-0), they lie higher than the respective theoretical values. Such a dispersive character of the energy migration can be explained qualitatively by the presence of fluorescent centers with 0-0 transitions differing from the “mean” at λ0-0.

Spectroscopic manifestations of flavomononucleotide dimers in polyvinyl alcohol films.

The present note comments on several publications which appeared in different journals containing many inaccurate statements and lacking honest citations of basic papers dealing with the application of solvatochromism to determine excited state dipole moments.

Excitation energy transport between the ionic forms of rhodamine B in viscous solutions

A new method for extending the utilizable range of Förster resonance energy transfer (FRET) is proposed and tested by the Monte Carlo technique. The obtained results indicate that the efficiency of FRET can be significantly enhanced at a given distance if the energy transfer takes place toward multiple acceptors that are closely located on a macromolecule instead of a single acceptor molecule as it is currently used in FRET analysis. On the other hand, reasonable FRET efficiency can be obtained at significantly longer distances than in the case of a single acceptor. Randomly distributed and parallel orientated acceptor transition moments with respect to the transition moment of the donor molecule have been analyzed as two extreme cases. As expected, a parallel orientation of donor and acceptor transition moments results in a more efficient excitation energy transfer. This finding could be used to directly reveal the assembly/deassembly of large protein complexes in a cell by fluorescence microscopy.