The factors limiting micropollutant biodegradation in the environment and how to stimulate this process have often been investigated. However, little information is available on the capacity of microbial communities to retain micropollutant biodegradation capacity in the absence of micropollutants or to reactivate micropollutant biodegradation in systems with fluctuating micropollutant concentrations. This study investigated how a period of 2 months without the addition of micropollutants and other organic carbon affected micropollutant biodegradation by a micropollutant-degrading microbial community. Stimulation of micropollutant biodegradation was performed by adding different types of dissolved organic carbon (DOC)—extracted from natural sources and acetate—increasing 10 × the micropollutant concentration, and inoculating with activated sludge. The results show that the capacity to biodegrade 3 micropollutants was permanently lost. However, the biodegradation activity of 2,4-D, antipyrine, chloridazon, and its metabolites restarted when these micropollutants were re-added to the community. Threshold concentrations similar to those obtained before the period of no substrate addition were achieved, but biodegradation rates were lower for some compounds. Through the addition of high acetate concentrations (108 mg-C/L), gabapentin biodegradation activity was regained, but 2,4-D biodegradation capacity was lost. An increase of bentazon concentration from 50 to 500 µg/L was necessary for biodegradation to be reactivated. These results provide initial insights into the longevity of micropollutant biodegradation capacity in the absence of the substance and strategies for reactivating micropollutant biodegrading communities.Graphical abstract