Evaluation of fluorescein benzoate di/tri/tetrasulfides as potential chemical tools and probes for reactive sulfur species

Reactive sulfur species (RSS) play vital roles in biological redox regulation but remain challenging to detect selectively. In this work, we prepared and tested fluorescein benzoate disulfide (SPS2), trisulfide (SPS3), and tetrasulfide (SPS4) as possible fluorescent probes for RSS. All these compounds exhibited good stability in aqueous buffers. The disulfide compound (SPS2) was found to be least reactive, and it only reacted with hydrogen disulfide (H 2 S 2 ) to give fluorescence. The trisulfide and tetrasulfide compounds (SPS3 and SPS4) were more reactive and they reacted with all nucleophilic RSS, including H 2 S 2 , H 2 S, and biothiols to show fluorescence. These results provide valuable insights into the properties of symmetric di-, tri-, and tetrasulfide structures under biologically relevant conditions. • A series of symmetric fluorescein benzoate di/tri/tetrasulfides (SPS2, SPS3, SPS4) were prepared and tested as probes for reactive sulfur species. • The disulfide compound SPS2 exhibited high selectivity toward H 2 S 2 and was successfully applied in cell imaging studies. • The tri- and tetrasulfides (SPS3 and SPS4) exhibited excellent stability and general reactivity toward all nucleophilic sulfur species including H 2 S, H 2 S 2 , and biothiols.