engleski

Mathematical model for fluorescence emission properties of PRODAN in different ethanol solutions

The molecular probe 6-propionyl-2-dimethylaminonaphthalene, PRODAN, is the choice for membrane and biomolecular structures investigations because of a strong dependence of its fluorescence on the environmental polarity. However, for the unequivocal interpretation of the results gained by such experiments the behavior of the probe molecule in different environmental characteristic must be known. Several detailed studies upon PRODAN fluorescence behavior have been preformed and theory argued since it has been designed but the unanimous interpretation is not yet accepted. Our aim was to determine the reliable method for resolving the complex spectra of PRODAN in pure solvents of different ethanol concentrations and to establish the sensitivity of the method for the interpretation of detected changes in PRODAN spectra when it is in complex biological structures. The samples of PRODAN were prepared in solutions of ethanol in 0.01 M phosphate buffer with 0.2 M NaCl. The final PRODAN concentrations of 0.25 ; 0.5 ; 1.5 ; 2 and 3 microM were measured in samples with ethanol concentrations of 0.3 ; 0.9 and 3 M. The spectra in pure buffer solution and in 100% ethanol solution were measured to support the discussion about environmental polarity importance. The steady state fluorescence spectra at two temperatures 25˚ C and 37˚ C were recorded on Carry Eclipse, Varian spectrofluorimeter at the Laboratory for Microwave spectroscopy, R. Bošković Institute, Zagreb, Croatia. The refractive indexes of the ethanol solutions have been determined by standard refractometer at temperature of 25˚ C. The dielectric responses of all investigated solutions at 20˚ C were measured by usage of home-made capacitive chamber in conjunction with Agilent 4294A precision impedance analyzer from 40 to 100 MHz at temperature of 25˚ C at the Institute of Physics, Zagreb. The corrected steady state PRODAN spectra showed complex structure. The model of two fluorescence transitions has been applied in analysis of those spectra. The experimental data have been fitted with the sum of two Gaussian functions: where A1, A2 and       are maximal fluorescence intensities and corresponding wavelengths of the two transitions respectively.   and    are related to curves halfwidth      and      by                       The behavior of two transition curves has been explained by differences in the pronounced solvent dipolar relaxation phenomena and consequently the existence of excited states with different energies depending on media polarity.

PRODAN; ethanol solutions; mathematical model

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