Introduction to the fluorescence spectroscopy principle
A molecule upon interacting with incident light may scatter a part of the light ( elastically, e.g., Rayleigh scattering or inelastically, e.g., Raman scattering) and may absorb rest of the light. An absorption of light may occur when the photon energy is equal to the energy difference between two energy states of the molecule. The interaction of the oscillating electromagnetic field of the radiation with the charged particles (electrons) in the molecule causes absorption of light. The absorption of light energy places the molecule in one of its many possible higher energy (excited) rotational, vibrational or electronic states, depending on the amount of the absorbed energy. Thus the molecule undergoes a transition to an upper energy or excited state. This is known as excitation or absorption process. The absorption process is extremely fast (takes approx. 10-15 s). All electronically excited states have a finite lifetime during which the excited state equilibrates with its surroundings. Therefore, after reaching the excited electronic state, the molecule returns to its ground state by losing the absorbed energy via various pathways that are either radiationless, in which no photons are emitted (energy converted into the disordered thermal motion of its surroundings), or radiative decay, which involves the emission of a photon. For example, the molecule can lose the energy by internal conversion (heat), quenching (external conversion), by emission of a photon (fluorescence), or by first intersystem crossing and then emission of a photon (phosphorescence). A schematic of the processes that occur following the electronic transition is given by ‘Jablonski diagram’ after Polish physicist, Aleksander Jablonski.