Coral reef fishes were recently discovered to have ultraviolet (UV) radiation screening compounds, most commonly known as mycosporine-like amino acids (MAAs), in their external body mucus. However, little is known about the identity and quantity of MAAs in the mucus of reef fishes or what factors affect their abundance and distribution. Using spectrophotometry, we examined the relative field UV absorbance of fresh mucus and quantified the UV absorbance per mg of mucus using laboratory UV absorbance for 7 species of reef fishes (Labroides dimidiatus and Thalassoma lunare [Labridae]; Chlorurus sordidus, Scarus flavipectoralis, S. niger, S. rivulatus, and S. schlegelii [Scaridae]) from Queensland, Australia. For the first time in fish mucus, we identified and quantified MAAs using High Performance Liquid Chromatography. In addition, to examine geographical variation in MAAs, MAA absorbance in T. lunare mucus was compared among 3 different locations (Lizard Island, Heron Island, and North Stradbroke Island) on the coast of Queensland. The mucus of all fish investigated contained MAAs. Depending on the species, different combinations and quantities of the MAAs asterina-330, palythene and mycosporine-N-methylamine serine were present. UV absorbance and MAA levels in mucus of T. lunare from North Stradbroke Island were lower (mycosporine-N-methylamine serine) or absent (palythene) compared with mucus from Lizard Island and Heron Island. This study shows that the identity and quantity of MAAs in fish mucus not only varies among species but also within a species sampled in different locations. This suggests that the ecological role of MAAs in fishes is varied and complex and that several factors may affect the type and quantity of MAAs found in the mucus of reef fishes.