Tunable Triazole-Dansyl Fluorescent Probes for Glutathione Detection: Modulating Probe-Biomarker Interactions for Enhanced Sensitivity and Selectivity in Biological, Environmental, and Cellular Systems.

A structurally tunable library of six triazole-dansyl fluorophores was investigated for substitution-dependent recognition of glutathione (GSH). All probes showed fluorescence quenching upon GSH addition, with the aliphatic-aliphatic derivative P4 exhibiting the highest sensitivity (LOD: 123 nM). High selectivity toward GSH was observed over competing amino acids, metal ions, and biomolecules, with minimal interference under competitive conditions. Mechanistic studies (UV-Vis, ESI-MS, 1H NMR, FTIR, PXRD, molecular docking, and DFT) revealed multivalent noncovalent interactions with a 2:1 binding stoichiometry, inducing perturbation of the intramolecular charge transfer (ICT) state. Time-resolved fluorescence confirmed a dynamic quenching pathway, while quantum yield measurements showed up to 49% quenching. pH- and temperature-dependent studies established optimal performance under physiological conditions. Practical applicability was demonstrated in human serum, cow urine, and goat blood with recoveries of 95-115% and negligible matrix effects. P4 did not exhibit any significant cytotoxicity in HeLa cells even at 100 μM. Fluorescence-based detection of GSH in HeLa cells and A549 cells confirmed the probe's sensing capability in mammalian cells, while cellular imaging established its suitability for bioimaging applications.