In the flavin family of photoactive biomolecules, lumichrome (LM) is a very important compound. It contains a tri-cyclic structure with methyl groups at two sides. It formed by the partial decomposition and biodegradation of riboflavin in both acidic as well as in neutral medium. Herein, we have studied the photophysical properties of LM in the presence of two zwitterionic surfactants, namely dodecyldimethyl(3-sulfopropyl) ammonium hydroxide inner salt (DSB), and tetradecyldimethyl(3-sulfopropyl) ammonium hydroxide inner salt (TSB), having the same head group but a different tail part. We have used steady-state absorption, fluorescence emission, and time-resolved fluorescence emission measurements. We observed that in the presence of zwitterionic surfactant aggregates LM shows excitation and emission wavelength dependent emission properties, which demonstrate the structural changes that take place from one form to another prototropic form of LM molecule. The higher rotational relaxation time of LM in the case of DSB compared to TSB demonstrated that LM is facing more rigid environment in DSB micelles compared to TSB micelles.