The absorption and fluorescence spectra of 3-hydroxyflavone (3-HF) in different polar solvents were observed with UV-Vis spectrometer and fluorescence spectrometer, respectively. There are three absorption bands in the absorption spectra, wherein two absorption bands with absorption peak at 300 and 345 nm, respectively, are strong, and the other one with absorption peak at 415 nm is weak. When the samples in different polar solvents were excited by 345 nm light, there appeared two new fluorescence bands peaked at 400 and 526 nm, respectively. The fluorescence band at 400 nm is attributable to the emission from enol structure and its intensity increases with increasing the polarity of protic solvents; that at 526 nm is attributable to the emission from the isomer structure and its intensity decreases with increasing the polarity of protic solvents. The results show that the increase in the polarity of protic solvents prevents the formation of isomer. When the samples in different polar solvents were excited by 415 nm light, three new fluorescence bands peaked at 440, 471 and 515 nm have not been reported so far. In order to identify the three new fluorescence bands, we prepared the samples with pH value of 5.0, 4.0 and 3.0 through incorporating the different amounts of acetic acid into 3-HF solution. The fluorescence spectra in different pH value solution were observed under excitation of 415 nm light, and it was found that the intensity of two fluorescence bands in the region of shorter wavelength changes with pH values changing. For identifying the fluorescence band of 515 nm peak wavelength, we put sodium hydroxide into 3-HF in ethanol solution and prepared 3-HF samples with pH values of 8.0, 8.5, 9.0, and 10.0. When the samples were excited by the 415 nm light, it was found that two fluorescence bands in the region of shorter wavelength disappeared and the intensity of the fluorescence band of 515 nm was enhanced. Since in sodium hydroxide solution 3-HF forms anion easily, we ascribed the fluorescence band with 515 nm peak wavelength to the emission from the 3-HF anion.