Excited-state intermolecular proton transfer of D-luciferin in reverse micelles has been investigated using steady-state and time-resolved fluorescence spectroscopy measurement. The different polar cores have been chosen for the study of proton transfer dynamics in aerosol-OT (AOT) reverse micelles. It is shown that aqueous reverse micelle is the suitable environment for the photoprotolytic reaction of D-luciferin. The neutral form of the chromophore is present both in ground and excited state at W-0 = 0. The proton transfer in nanometer size water pool of water/AOT/n-heptane begins at W-0 = 8 and increases with increasing W-0 values. However, the intermolecular excited-state proton transfer (ESPT) of D-luciferin is inhibited in nonaquous reverse micelles with DMF and DMSO as a polar core. Thus, the requirement of ESPT of D-luciferin to take place in reverse micelles consists of polar protic solvent like water as a polar core.