Two types of luminescence observed from proteins are fluorescence and phosphorescence. Fluorescence arises from the decay of the excited single state S1 to the ground state S0. Phosphorescence arises from the decay of the excited triplet state T1 to the ground state S0. Phosphorescence is strictly forbidden by quantum mechanics since it involves a transition between pure states of different spin multiplicity, however, this forbiddenness can be relaxed by interactions between the magnetic dipoles generated by the spin of the electron and the orbital motion of the electron. These spin-orbit interactions lead to coupling of the singlet and triplet states, giving some singlet character to the triplet state, and thus to a small, but finite, transition probability, between T1 and S0.

Spin-orbit coupling is weak in planar, aromatic hydrocarbons like tryptophan so other perturbations must also be included to explain the long phosphorescence lifetime observed. The important perturbation is spin-vibronic coupling which couples together nuclear vibrational motions to the electronic states giving rise to further mixing of singlet and triplet states.

Molecular oxygen is an excellent quencher of phosphorescence and so room temperature phosphorescence (RTP) from tryptophans in proteins was not discovered until the samples were thoroughly deoxygenated². (Proteins were observed to phosphoresce at only low temperature before this time). Since that discovery was made, rtp has been detected in many proteins^1,3,4 and has been shown to be a useful tool for studying protein dynamics and stability.