Simultaneous Detection of Hydrogen Sulfide and Peroxynitrite Dysregulation with a Sequentially Activated Fluorescent Probe in Ulcerative Colitis Disease

Ulcerative colitis (UC), often referred to as "green cancer", is a chronic inflammatory bowel disease with an unclear etiology, closely associated with the imbalance of hydrogen sulfide (H₂S) and peroxynitrite (ONOO^-). H₂S exhibits anti-inflammatory effects at physiological levels, but excessive concentrations can compromise the intestinal barrier, while ONOO^- aggravates inflammation. To facilitate the molecular-level monitoring of these compounds in UC, we developed a novel fluorescent probe, BCH, capable of simultaneously detecting H₂S and ONOO^- via distinct fluorescent channels in a cascade mode. BCH features rapid response times (H₂S: 6 min; ONOO^-: 5 min) with well-separated emission peaks (120 nm), minimizing spectral overlap. The probe demonstrates high selectivity and good detection linearity for H₂S (0-500 μM) and ONOO^- (0-100 μM), with detection limits of 1.6 μM for H₂S and 0.05 μM for ONOO^-. BCH has been successfully applied for imaging H₂S and ONOO^- in living cells and zebrafish, enabling the sensitive detection of their fluctuations in a UC mouse model. These findings indicate that BCH is highly promising for studying the roles of H₂S and ONOO^- in UC and assessing their biological functions.