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Most information on the occurrences of active pharmaceutical ingredients (APIs) in natural water bodies comes from developed nations, leaving developing countries like Ethiopia understudied. This study examines the presence and potential ecological risks of APIs in Lake Hawassa, a tropical freshwater lake in the Ethiopian Rift Valley. The study analyzed 36 APIs from 11 therapeutic classes of human medications in water and sediment samples. The risk quotient (RQ) method was used to assess ecological risks to aquatic organisms. Forty-five composite samples of water and sediment were collected from five sites during three sampling events. The samples were analyzed in the Chemistry laboratory at the University of Umeå, Sweden using the ultrahigh-performance liquid chromatography mass spectrometer (UHPLC-MS/MS) method. The results indicated that 16 APIs were detected in the water samples and three in the sediment. Ciprofloxacin, artesunate, mefloquine, fluconazole, and tramadol were the most frequent APIs detected in the analyzed water samples. The concentration of ciprofloxacin was the highest of all the APIs, detected with RQ > 1, indicating high risk to the aquatic biota in the Lake Hawassa ecosystem. Similarly, diclofenac, and sulfonamide in water, lorazepam, mefloquine, and lumefantrine in sediments, occurred at concentrations that could cause a medium risk (RQ > 0.1) to the aquatic biota. The occurrence of the APIs in the Lake Hawassa ecosystem affirms early warning, and the Antibiotics may create Antibiotic-resistant genes posing a public health threat. Therefore, it is crucial to implement systems for gathering unused medications and monitoring waste disposal and sewage treatment in the study area. Despite the limited research on APIs in the region where this study was conducted, our findings are highly valuable for understanding APIs in freshwater ecosystems worldwide. Our results align with global concerns about APIs, making them a valuable tool for creating universally applicable guidelines.