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Organisms exposed to suboptimal conditions face an a priori cost of combating stress in terms of metabolic resources. The energy available for maintenance, growth, and reproduction, based on the biochemical analysis of the energy budget rather than on the direct measurement of those endpoints, may therefore provide a sensitive measure of stress in an organism. The aim of this study was to validate changes in energy budget as an ecologically relevant sublethal parameter by comparing these responses with physiological, growth, and reproductive endpoints. A toxicity test with zebrafish (Danio rerio) was conducted where fish were exposed to a control and 50, 75, and 100% effluent for 28 d under flow‐through conditions. Effects of effluent exposure were measured as changes in glycogen, lipid, and protein budgets. Furthermore, the observed effects were linked to different processes like growth, condition …