The low cost and free sale of some medicines are two of the main factors behind the considerable increase in the consumption of non-steroidal anti-inflammatory drugs around the world, particularly ketoprofen (Kp). Despite its widespread use, the ecotoxicological profile of Kp remains poorly explored and therefore, to date, the intra- and interspecific effects of Kp have yet to be explored. Therefore, this study aimed to provide a classification of the environmental hazard of Kp, firstly by determining the intraspecific tolerance of a laboratory-simulated cladoceran population comprising five different clonal lineages selected accordingly to their previous tolerance to copper contamination; and secondly, by providing the lethal and sublethal concentrations for effect (LC_x and EC_x, respectively) of Kp for freshwater species representative of the different trophic levels. The LC_x were later assembled into Species Sensitivity Distribution (SSD) curves to derive hazard concentrations that affect 5% of the population (HC₅, probabilistic approach) and compared with the deterministic approach (lowest L(E)_x divided by an assessment factor of 1000). Risk quotient for Kp was determined based on the most conservative value. In relation to the results with the simulated population in the laboratory with clones of Daphnia longispina, the results showed significant correlation (0.97) between their baseline tolerance to copper and their tolerance to a compound they had never been exposed to before, such as ketoprofen. This type of approach, which is generally neglected in ecological risk assessment, can help to outline more realistic environmental protection targets, since genetic diversity is the cornerstone of population resilience in the face of environmental change. Regarding interspecific variability, the results showed that the embryos of the fish Danio rerio were the most sensitive ecological receptors, while the green microalga Raphidocelis subcapitata was the most tolerant to Kp. The estimated LC₅₀ values (95% confidence limits) were 6.16 (5.17 - 7.43) mg Kp/L and 47.2 (28.3 - 66.7) mg Kp/L for D. rerio and R. subcapitata, respectively. The estimated HC₅ of the SSD was 4.91 mg Kp/L. Determination of the risk quotient [based on the lowest and most conservative L(E)C₅₀/1000] indicated a quotient >1, meaning that Kp poses a risk to aquatic biota. Given that ketoprofen is an increasingly used drug and has the potential to have long-lasting effects, its ecotoxicity deserves to be further explored considering various exposure scenarios.