coelenterazine and dioxetanone

Imidazopyrazinone is a typical scaffold present in marine bioluminescence, in which thermal energy is converted into excitation energy in an enzyme-catalyzed reaction. In fact, the imidazopyrazinone scaffold is a common link among organisms of eight phyla. The characterization of the light emission mechanism is essential for the development of future applications in bioimaging, bioanalysis and biomedicine. Herein, we have studied the chemiluminescent reaction of three commercially-available imidazopyrazinones (Cypridina luciferin, Coelenterazine and Coelenterazine-e) in several aprotic solvents at different pH. We have found that at acidic pH only DMF and DMSO consistently present high light emission, while chemiluminescence in other solvents is negligible. We have attributed this to the inability of most solvents to allow for the deprotonation of the imidazopyrazinone core, thereby preventing the oxygenation step. We have also observed that increasing the pH of the solution leads to the inhibition of chemiluminescence, which we attributed to the deprotonation of the dioxetanone intermediate, as the neutral species is the one associated with efficient chemiexcitation. We have also observed that the pK

Topics: Heterocyclic Compounds, 1-Ring; Hydrogen-Ion Concentration; Imidazoles; Luciferases; Luminescence; Luminescent Agents; Pyrazines; Solvents

Coelenterazine is a widespread bioluminescent substrate for a diverse set of marine species. Moreover, its imidazopyrazinone core is present in eight phyla of bioluminescent organisms. Given their very attractive intrinsic properties, these bioluminescent systems have been used in bioimaging, photodynamic therapy of cancer, as gene reporter and in sensing applications, among others. While it is known that bioluminescence results from the thermolysis of high-energy dioxetanones, the mechanism and dioxetanone species responsible for the singlet chemiexcitation of Coelenterazine are not fully understood. The theoretical characterization of the reactions of model Coelenterazine dioxetanones showed that efficient chemiexcitation is caused by a neutral dioxetanone with limited electron and charge transfer, by accessing a region of the PES where ground and excited states are nearly-degenerated. This finding was supported by calculation of equilibrium constants, which showed that only neutral dioxetanone is present in conditions associated with bioluminescence. Moreover, while cationic amino-acids easily protonate amide dioxetanone, anionic ones cannot deprotonate the neutral species. These results indicate that, contrary to existent theories, efficient chemiexcitation can occur with significant electron and/or charge transfer. In fact, these processes can be prejudicial to chemiexcitation, as anionic dioxetanones showed a less efficient chemiexcitation despite the occurrence of significant electron and charge transfer.

Topics: Amides; Electron Transport; Heterocyclic Compounds, 1-Ring; Imidazoles; Luminescence; Models, Chemical; Models, Molecular; Molecular Conformation; Pyrazines