Development of a Rapid-Response Fluorescent Probe for H2S: Mechanism Elucidation and Biological Applications

Hydrogen sulfide (H₂S) is an important signaling molecule involved in various physiological and pathological processes, making its accurate detection in biological systems highly desirable. In this study, two fluorescent probes (<b>M1</b> and <b>M2</b>) based on 1,8-naphthalimide were developed for H₂S detection via a nucleophilic aromatic substitution. <b>M1</b> demonstrated high sensitivity and selectivity for H₂S in aqueous media, with a detection limit of 0.64 µM and a strong linear fluorescence response in the range of 0-22 µM of NaHS. The reaction kinetics revealed a rapid response, with a reaction rate constant of 7.56 × 10² M^-1 s^-1, and <b>M1</b> was most effective in the pH range of 6-10. Mechanism studies using ¹H NMR titration confirmed the formation of 4-hydroxyphenyl-1,8-naphthalimide as the product of H₂S-triggered nucleophilic substitution. <b>M1</b> was applied in MDA-MB-231 cells for cell imaging, in which <b>M1</b> provided significant fluorescence enhancement upon NaHS treatment, confirming its applicability for detecting H₂S in biological environments. In comparison, <b>M2</b>, designed with extended conjugation for red-shifted emission, exhibited weaker sensitivity due to the reduced stability of its naphtholate product and lower solubility. These results demonstrate that <b>M1</b> is a highly effective and selective fluorescent probe for detecting H₂S, providing a valuable resource for investigating the biological roles of H₂S in health and disease.