The consumption of synthetic detergents is increasing year by year due to increasing urbanization, which reflects on higher concentration of this pollutant in the environment. In order to purify wastewaters from different pollutants, the application of new technologies such as bioremediation is necessary. From the environmental point of view, it is important to identify microorganisms that are tolerant to the presence of this pollutant. This chapter presents the experimental evaluation of ability of several fungal species, isolated from municipal sewage and industrial wastewater, in removing a high detergent concentration (anionic surfactants) from the environment as well as their potential application in biotechnology. The selected fungi were cultivated in Czapek-Dox liquid medium supplemented with commercial detergent "Merix" (Henkel, Kruševac, Serbia). Changes of physicochemical and biochemical parameters such as pH, redox potential, dry weight biomass, and enzymes activities such as alkaline protease and phosphatase were evaluated during 16 days of cultivation. The obtained results could be useful in the implementation of tested fungi in bioremediation processes and in biotechnology.