The crystal structure of the new polymorphic form of 3-aminoflavone (3-AF) has been determined by single crystal X-ray diffraction. This report presents results of fluorimetric studies on 3-AF in methanol and aquatic solvents. Based on 3D fluorescence emission spectra, optimal values for excitation (λex) and emission/analytical (λem) wavelength, the analytical concentration range as well as the range of concentration quenching for the studied compound were established. Moreover, the limit of detection (LOD) and the limit of quantification (LOQ) were determined. The results were compared with those obtained using the standard UV-Vis absorption spectrophotometric method. The effect of acidity (pH) and the concentration of halide anions (chlorides, bromides, iodides and fluorides) on fluorescence quenching were analysed.