The behavior and stability of dissolved reduced glutathione (GSH), an environmental antioxidant and metal transporter, is poorly known in natural waters. Glutathione oxidation rates were determined in both fresh- and brackish waters artificially submitted to different radiation wavebands. Photo-oxidation of GSH followed pseudo-first order kinetics, with half-lives ranging from 4 to 30h in natural freshwater, and from 1.6 to 7h in saltwater, with generally shorter persistence in UV-irradiated surface waters than in dark treatments. Chloride was shown to indirectly promote GSH photo-oxidation, probably through its role in the formation of radicals. The addition of Cu(II) to synthetic waters resulted in the rapid oxidation of GSH. The addition of Hg(II), a metal with strong affinity for thiols, protected GSH from oxidation by Cu(II) in the dark, but not under UV radiation. We conclude that UV-induced photo-oxidation is a key process altering the fate of GSH in natural waters. Also, the formation of stable GSH–Hg complexes could increase the bioavailability of Hg towards microorganisms in aquatic systems.