Pyrene fluorescence fine structure as a polarity probe of hydrophobic regions: Behavior in model solvents

The vibrational fine structure superimposed on pyrene monomer fluorescence is solvent dependent at room temperature. The fine structure pattern of pyrene monomer fluorescence was found to be independent of excitation conditions or collisional quenching, but highly dependent on solvent polarity. Variation in pyrene fluorescence fine structure was investigated in alkane and alkanol solvents and solvent mixtures. Although changes in the fine structure pattern were detected across the monomer fluorescence spectrum, the greatest difference occurred at the 373- and 384-nm peaks. In nonpolar environments, the 384-nm peak dominated the pyrene spectrum, whereas at higher polarity, the 373-nm peak was the most intense. By expressing the relative emission intensities at 373- and 384-nm peaks as a ratio, a sensitive parameter of solvent polarity was obtained. The $\text{373}\text{384}$ peak ratio results yielded a curvilinear relationship as a function of dielectric constant. This effect can be applied to monitor changes in polarity of pyrene microenvironments in proteins and membranes.