The stoichiometry and apparent stability constants (beta) of the complexes formed between Ni2+ and four different hydroxyflavones (3-hydroxyflavone, 5-hydroxyflavone, chrysin, and galangin) were determined in methanolic solutions. A multivariate curve resolution methodology was applied to estimate the beta values using UV-vis spectroscopic data. All the complexes exhibit 1:2 metal:ligand stoichiometry. The highest and lowest beta values were obtained for the galangin and 5-hydroxyflavone complexes, respectively. The formation of the complexes was confirmed by Fourier transform infrared spectroscopy. Time-dependent density functional theory and natural transition orbital analysis were performed to describe the spectroscopic features of the studied compounds, and quantum theory of atoms in molecules was applied to evaluate different intermolecular interactions. Good correlations were obtained between the calculated UV-vis absorption spectra using the M06 functional and the experimental ones. The main absorption band of these Ni2+ complexes have an important metal-to-ligand charge transfer mixed with an intraligand charge transfer (ILCT). Two possible chelation sites were considered for the interaction of Ni2+ with galangin. The combination of spectroscopic and quantum chemistry calculations results indicate the existence of an equilibrium between the two carbonyl-hydroxyl sites of galangin in the Ni2+ complex formation.