Quenching of acridine orange fluorescence by salts in aqueous solutions: Effects of aggregation and charge transfer

Acridine orange (AO) is widely applied in biology and medicine as a fluorescence probe, an intracellular pH indicator, and a photosensitizer in photodynamic therapy due to its adequate spectroscopic characteristics and high affinity to biological structures. Being introduced in an organism, AO is dispersed in blood plasma characterized by high ionic strength (ca. 0.36 M in humans). We have investigated the effect of ionic strength upon AO spectral characteristics and fluorescence quenching. The effect of pH on these characteristics was also tested. Salts quench AO fluorescence, the quenching constant (kq) increasing with the AO concentration. Salts stimulate AO aggregation, the process depending weakly on the salt origin. On the other hand, kq does depend on the salt anion origin, increasing as the anion oxidation potential decreases, and is virtually independent of the cation origin. This means that at least two different mechanisms of the AO fluorescence quenching by salts exist: fluorescence intensity decrease due to AO aggregation and quenching by partial electron transfer from salt anion to AO molecule in its singlet excited state (the exciplex formation).