Fluorescence Spectroscopy

Study Notes

In fluorescence spectroscopy, a sample is irradiated with monochromatic radiation of a certain wavelength (excitation), and a scan of the wavelength of emitted radiation is recorded.
The excitation and emission spectra of a compound are plotted on the same chart, showing a displacement of the emission band to a longer wavelength (Stock's shift) and a mirror image relationship between the two spectra.

The quantum yield or quantum efficiency (f) for a fluorescent process is the ratio of the number of molecules that fluoresce to the total number of excited molecules or the ratio of the number of photons emitted to that absorbed.
For highly fluorescent molecules, f approaches unity (f = 1), while for non-fluorescent molecules, f = 0.

Fluorescence intensity (F) is proportional to concentration (c) and can be calculated using the equation: F = 2.3 K εbc I0 or F = K`c.
A plot of fluorescence intensity versus concentration is linear at low concentrations, but linearity is lost at high concentrations due to self-absorption and self-quenching.

Molecular structure: Compounds containing aromatic functional groups, aliphatic and alicyclic carbonyl groups, or conjugated double-bond structures can exhibit fluorescence.
Temperature and solvent: Increasing temperature and decreasing solvent viscosity can decrease fluorescence, while polar solvents can enhance fluorescence.
Dissolved oxygen: Oxygen can quench fluorescence by accepting energy from excited molecules.

Phosphorescence is rare in solution at room temperature due to triplet states being susceptible to collisions with solvent molecules and quenching by oxygen.
Phosphorescence can be observed by dispersing the sample in a rigid matrix or freezing the solution to a low temperature.

Molecules with fused rings, such as fluorene, are especially fluorescent, and the extent of fluorescence is directly proportional to the number of rings in the molecule.
Halogen substitution, especially with bromine and iodine, can decrease fluorescence due to intersystem crossing.
Rigid planar structures, such as in fluorene, can favor fluorescence, while non-rigid molecules tend to lose energy through non-radiative means and do not fluoresce much.