New 9H-pyrido[2,3-b]Indoles, Synthesis, Photophysical/Theoretical Studies, and Application as the Fluorescence Probes

Using the "1,2,4-triazine" methodology, we have designed a series of new 9H-pyrido[2,3-b]indole-based fluorophores/probes 4a-e, 7a,b, and 8a,b. Photophysical studies of the push-pull fluorophores 8a and 8b revealed a positive solvatochromic effect with Δμ > 15D and Stokes shifts of up to 270 nm. Experimental data and theoretical calculations of the ground and excited states of the compounds demonstrated that pyridoindoles 4a-e and 7a,b exhibit a planarized intramolecular charge-transfer (PLICT) state, which is accompanied by a gradual increase in luminescence quantum yield for samples 4d and 7b with increasing solvent polarity. Probe 4b, characterized by an acid dissociation constant (pK_a) of 5.5, showed high sensitivity to local pH changes in the microenvironment. A "turn-off" acidochromic response was observed for 4d and 8a with Stern-Volmer quenching constants of 1.6 × 10³ and 5.5 × 10³ M^-1, respectively, while a "turn-off-on" response was detected for 4b and 8b during titration with TFA. Fluorescence and UV-vis absorption spectroscopy studies of the interaction between probe 4b and calf thymus DNA (ctDNA) confirmed a static quenching mechanism with the formation of 4b-ctDNA complex, indicating that the probe binds to the DNA helix via a groove binding mode. These findings highlight the potential of this pyridoindoles for visual detection and cellular imaging applications.