The characteristic fluorescence properties of quercetin-3-O-rhamnoside (QCRM) and quercetin-3-O-rutinoside (QCRT) were studied in CH3OH–H2O and CH3CN–H2O mixed solvents. Although QCRM and QCRT are known as nonfluorescent molecules, significant fluorescence emissions were discovered at 360 nm in CH3OH and CH3CN when they were promoted to the second excited state. The emission band is broad and structureless and the intensity decreases quickly as the H2O composition in the solvent increases. When the amount of H2O exceeds 60% in both mixed solvents, this emission disappears due to the formation of the distorted excited state. This state will be formed due to the strong intermolecular hydrogen bonding between the polar groups of solute and H2O. As the composition of CH3OH or CH3CN in solvent becomes large, the number of molecules having several intramolecular hydrogen bonding increases. Some of these molecules will be changed to a fluorescent species during the decay process, after excitation. The theoretical calculation further supports these results. The change of the lifetimes, quantum yields, and radiative and nonradiative rate constants of molecules was also examined as a function of solvatochromic parameters for CH3OH–H2O and CH3CN–H2O.