Cephalexin (CEX) is an antibiotic commonly used to treat bacterial infections in humans and animals. However, it is also a micropollutant. Thus, this study evaluated the degradation of CEX using ultraviolet irradiation (UV-C) and analyzed the by-products as well as their residual antimicrobial activity. A reactor with a mercury vapor lamp was used for the degradation. Irradiated CEX solutions were collected over a period of 4 hours and analyzed using high-performance liquid chromatography coupled with mass spectrometry. For the residual antimicrobial activity the susceptibility test was performed using Staphylococcus aureus and Escherichia coli microorganisms by broth microdilution. It was found that CEX, after treatment, generated a metabolite with a mass of 150 m/z in 15 min. A four- and eightfold increase in the minimum inhibitory concentration of the drug against S. aureus and E. coli could be observed, respectively, after 20 min. Therefore, this treatment proved to be effective in the degradation of CEX, being able to degrade 81% of the initial molecule of the drug in 20 min. Furthermore, the antimicrobial activity of the CEX solution decreased as the irradiation time increased, indicating loss of antimicrobial function of the initial CEX molecule and the resulting by-products.