Natural Deep Eutectic Solvents (NADES) are composed of supramolecular interactions of two or more natural compounds, such as organic acids, sugars, and amino acids, and they are being used as a new media alternative to conventional solvents. In this study, a new application of NADES is presented as a possible technology for biofilm structural breaker in complex systems since the current solvents used for biofilm cleaning and extraction of biofilm components use hazardous solutions. The NADES (betaine:urea:water and lactic acid:glucose:water) were analyzed before and after the biofilm treatment by attenuated total reflection Fourier-transform infrared spectroscopy and fluorescence excitation-emission matrix spectroscopy. Our results indicate that the green solvents could solubilize up to ≈70 percent of the main components of the biofilms extracellular matrix. The solubilization of the biomolecules weakened the biofilm structure, which could enhance the biofilm solubilization and removal. The NADES have the potential to be an environment-friendly, green solvent to extract valuable compounds and break the main structure from the biofilm, leading to a greener method for extracellular polymeric substance (EPS) extraction and biofilm treatment in various water treatment systems.@en