The effect of solvent on the photoluminescence behavior of luminol was studied by steady-state fluorescence spectroscopy. The fluorescence spectral behavior of luminol is markedly different in polar protic solvents compared to that in aprotic solvents. A quantitative estimation of the contribution from different solvatochromic parameters, like solvent polarizibility (pi*), hydrogen-bond donor (alpha), and hydrogen-bond acceptor (beta), was made using the linear free energy relationship based on the Kamlet-Taft equation. The analysis reveals that the hydrogen-bond-donating ability (acidity) of the solvent is the most important parameter to characterize the excited-state behavior of luminol. Quantum mechanical calculations using density functional theory (DFT) predict the most stable structure, out of several possible tautomeric conformers of luminol with varying degrees of hydration. In the excited state, charge localization at specific points of the luminol phthalhydrazide moiety causes the solvent to interact primarily through hydrogen-bond donation.