Average Extinction Coefficient for Re-absorption of Fluorescence Radiation through Uranin Solution

Abstract

THE absorption and the fluorescence spectra of fluorescein in alkali solution overlap considerably. This overlap introduces a certain amount of error in the measurement of intensity of fluorescence in so far as it is not directly proportional to the incident radiation absorbed by the solution. If a fluorescent quantum is emitted somewhere deep in the solution, then during its passage from its point of emission to a measuring device like a photocell, there is a chance that it will be repeatedly re-absorbed and re-emitted by the solution. The consequence of this process is that the average life-time of the excited molecule is increased¹, as each absorption gives a fresh lease of life of about 10⁻⁸ sec. The effect will be greater, the higher the concentration of the fluorescing solution. Since intensity of fluorescence is directly proportional to the life-time, τ, of the fluorescent molecule under a given set of conditions τ₁/τ₂ = F₁/F₂, the intensity as measured by the photocell will not be the true intensity. The effect has been explained in different ways^1–3.