Solvent and Environmental Effects

Abstract

Solvent polarity and the local environment have profound effects on the emission spectral properties of fluorophores. The effects of solvent polarity are one origin of the Stokes shift, which is one of the earliest observations in fluorescence. Emission spectra are easily measured, resulting in numerous publications on emission spectra of fluorophores in different solvents, and when bound to proteins, membranes, and nucleic acids. One common use of solvent effects is to determine the polarity of the probe binding site on the macromolecule. This is accomplished by comparison of the emission spectra and/or quantum yields when the flu-orophore is bound to the macromolecule or dissolved in solvents of different polarity. However, there are many additional instances where solvent effects are used. Suppose a fluorescent ligand binds to a protein or membrane. Binding is usually accompanied by spectral shift or change in quantum yield due to the different environment for the bound ligand. Alternatively, the ligand may induce a spectral shift in the intrinsic or extrinsic protein fluorescence. In either case the spectral changes can be used to measure the extent of binding. The effects of solvent and environment on fluorescence spectra are complex, and are due to several factors in addition to solvent polarity. The factors that affect fluorescence emission spectra and quantum yields include: Solvent polarity and viscosity Rate of solvent relaxation Probe conformational changes Rigidity of the local environment Internal charge transfer Proton transfer and excited state reactions Probe–probe interactions Changes in radiative and non-radiative decay rates