Photophysical processes of ethidium bromide (EB) in homogeneous solutions, micelles, and reverse micelles have been investigated. In acetonitrile, the fluorescence intensity and lifetime of EB are 6.3 +/- 0.3 times those in water and 1.25 +/- 0.1 times those in acetone. This is attributed to the weaker hydrogen-bond acceptor property of acetonitrile, compared to water and acetone. Addition of water to acetonitrile leads to a marked quenching of the EB emission, with a quenching constant of (1.7 +/- 0.3) x 10(7) M-1 s(-1). In aqueous solution, hydroxyl ion quenches EB emission more dramatically with a quenching constant of (4.4 +/- 0.4) x 10(10) M-1 s(-1). This is ascribed to the abstraction of the amino proton of the excited ethidium ion by the hydroxyl ion. Emission properties of EB are found to be good monitors for the micellization of an anionic micelle, sodium dodecyl sulfate (SDS). In AOT microemulsion, EB exhibits nearly (1.8 +/- 0.1)-fold emission enhancement relative to water. The emission properties of EB are found to be independent of the water-to-surfactant ratio, w(0). In AOT microemulsion when instead of water D2O is injected, a further 2.3 times emission enhancement is observed. However, in AOT microemulsion, the hydroxyl ion does not quench the EB emission, even when a highly alkaline aqueous solution of EB (pH = 12.6)is injected into the reverse micelle. It is proposed that the anionic surfactant, AOT, strongly attracts the ethidium cation to the ACT-water interface but expels the hydroxyl anion from the ACT-water interface to the water pool, and hence, the hydroxyl anion cannot access the ethidium cation.