<p>Sol-gel materials are well-known glassy matrices for hosting functional optical molecules. Fluorene-derived chromophores are commonly used as two-photon-absorber dyes for nonlinear optics applications; however, the optical properties of asymmetric fluorene derivatives are very sensitive to their surrounding environment. In this work, we studied two formulations of a hybrid sol-gel material host doped with a diphenylamine–fluorene–benzothiazole chromophore. The stage at which the chromophore was added to the sol-gel mixture was found to significantly affect the optical properties of the dye in the sol-gel glass. The addition of the chromophore after mixing the sol-gel precursors but before gelation yields a green glass with absorption and fluorescence spectra that are similar to the chromophore in solution, while the addition of the chromophore before hydrolysis of the sol-gel precursors yields an orange glass with a new low energy absorption band and redshifted fluorescence compared to the green glass. Studies of the chromophore in acidic solution and ultrafast transient absorption studies of the sol-gel glasses confirm that the addition of the chromophore before hydrolysis of the sol-gel precursors leads to protonation of the chromophore and formation of an excited state with enhanced intramolecular charge-transfer character. These results suggest that variations in the formulation process of sol-gel optical components could provide an additional route to fine-tune the photophysical properties of the doped organic chromophores in applications that benefit from charge-transfer excited states.</p><p></p>