A ratiometric fluorescent probe for cysteine and glutathione differentiation and its application for cysteine detection in foods

Cysteine (Cys) and glutathione (GSH) are crucial biothiols implicated in cellular redox balance, necessitating accurate detection methods. Herein, we developed a novel fluorescent probe based on the fluorescent dye pyronin (PYR), capable of monitoring both Cys and GSH using ratiometric fluorescence signals. The probe, named PYR-CG, where "C" stands for Cys and "G" stands for GSH, demonstrates selective detection capabilities for Cys and GSH. UV absorption and fluorescence emission spectra changes revealed distinct shifts upon interaction with Cys and GSH. Theoretical and experimental studies validated PYR-CG's responsiveness to Cys and GSH, elucidating its mechanism as a ratiometric probe. Notably, PYR-CG exhibited rapid response kinetics for Cys and GSH, with low detection limits of 88 nM and 164 nM, respectively. Importantly, PYR-CG demonstrated exceptional selectivity for Cys and GSH over interfering analytes. Screening of molecules with similar structures to Cys further affirmed PYR-CG's specificity, particularly towards N-terminal Cys in peptides. Furthermore, recovery experiments underscored PYR-CG's reliability for quantitative Cys detection in complex food samples. Our findings highlight the innovative design and practical applicability of PYR-CG, offering a valuable tool for sensitive and selective detection of Cys in diverse biological and food samples.