||Artificial light at night (ALAN) changes the natural rhythm of light and darkness and can impair the biorhythms of animals, for example the nocturnal melatonin production of vertebrates, which serves as a proxy for daily physiological rhythms. Freshwater fish are exposed to ALAN in large urban and suburban areas in the form of direct light or in the form of skyglow, a diffuse brightening of the night sky through the scattered light reflected by clouds, atmospheric molecules, and particles in the air. However, investigations on the sensitivity of melatonin production of fish towards low intensities of ALAN in the range of typical skyglow are rare. Therefore, we exposed Eurasian perch (Perca fluviatilis) to nocturnal illumination levels of 0.01 lx, 0.1 lx and 1 lx and a control group with dark nights and daylight intensities of 2900 lx in all groups. After ten days of exposure to the experimental conditions, tank water was non-invasively sampled every 3 h over a 24 h period and melatonin was measured by ELISA. Melatonin was gradually reduced in all treatments with increasing intensity of ALAN whereas rhythmicity was maintained in all treatment groups although at 1 lx not all evaluated parameters confirmed rhythmicity. These results show a high sensitivity of Eurasian perch towards ALAN indicating that low light intensities of 0.01 lx and 0.1 lx as they occur in urban and suburban areas in the form of skyglow can affect the physiology of Eurasian perch. Furthermore, we highlight how this may impact perch in their sensitivity towards lunar rhythms and the role of skyglow for biorhythms of temperate freshwater fish.