Given the particular importance of dye photostability for single-molecule investigations, fluorescence fluctuation spectroscopy, and laser-scanning microscopy, refined strategies were explored for enhancing the fluorescence signal by selectively quenching the first excited triplet state of the laser dye Rhodamine 123 (Rh123). The strategy is to quench the T, state by Dexter triplet energy transfer while undesired quenching of the singlet state via Forster or Dexter sin-let energy transfer and the generation of Free radicals through electron transfer should be avoided. Diphenylhexatrienes (DPHs) were tested in ethanol for their beneficial effects as a novel class of photoprotective agents using fluorescence correlation spectroscopy. A library of DPHs with electron-donating (dimethlyamino) and withdrawing substituents (e.g., trifluromethyl) was synthesized to optimize the electronic properties. Quantum chemical calculations, Optical spectroscopy, and cyclic voltammetry were used to determine the electronic properties. The computed T-1 emission energy of Rh123 and the T-1 excitation energies of all DPHs allow for exergonic triplet energy transfer to the quencher. The parent compound quenches the T-1 state of Rh123 nearly diffusion controlled (4.9 x 10(9) M-1 s(-1)). All electron-deficient DPHs significantly increase (3x) the fluorescence rate of RH23 by reducing the triplet state population and by avoiding (he Formation of other long-lived dark radical states. The quenching constants are reduced by more than a factor of 2, if substituents with increasing size or electronegativity are introduced. The beneficial effect of triplet quenching of substituted DPHs is governed by a delicate interplay of steric, electronic, and intermolecular coulombic effects.