During routine fluorescence analysis, the presence of dissolved oxygen in solutions can result in the dynamic quenching of a fluorophore's emission through collisional deexcitation of the fluorophore's excited state. In order to avoid this type of fluorescence quenching, dissolved oxygen is often removed from solutions by an inert gas purging procedure. However, the details and quantification of this purging process are often limited in fluorescence studies. In this work, standard 10 mm × 10 mm fluorescence cuvettes are filled with polyaromatic hydrocarbon (PAH) solutions in cyclohexane and purged using nitrogen gas, and the experimental purging parameters (nitrogen flow rate, amount of volatile solvent loss, and rate of oxygen removal) are measured and analyzed. For experimental conditions similar to those used in this study, we are able to provide useful guidelines for the deoxygenation of solutions, specifically the purge times required for solutions of fluorophores with various fluorescence lifetimes. Enhancement factors, or <i>F</i><sub>0</sub>/<i>F</i> values (the ratio of fluorescence intensity of a completely deoxygenated solution to the fluorescence intensity of an aerated solution), for chrysene, phenanthrene, naphthalene, and pyrene solutions in cyclohexane were found to be 3.61 ± 0.02, 4.17 ± 0.02, 7.63 ± 0.07, and 21.81 ± 0.35, respectively.
Vol. 3, Iss. 4 Virtual Journal for Biomedical Optics
Todd Pagano, Adam J. Biacchi, and Jonathan E. Kenny, "Nitrogen Gas Purging for the Deoxygenation of Polyaromatic Hydrocarbon Solutions in Cyclohexane for Routine Fluorescence Analysis," Appl. Spectrosc. 62, 333-336 (2008)