Biothiols are essential reactive sulfur species (RSS), which play crucial roles in various critical physiological activities. To clarify their complex correlations in signal transduction and metabolism pathways, methods to distinguish H 2 S, cysteine (Cys), homocysteine (Hcy), and glutathione (GSH) at the same time are highly needed. Herein, we deliberately integrate 7-diethylcoumarin and resorufin through an ether bond to develop RC for simultaneous differentiation of Cys, Hcy, GSH, and H 2 S for the first time. RC displays distinct fluorescence colors in response to H 2 S, Cys, Hcy, and GSH as red, blue-red, bluegreen-red, and green-red, respectively, both in aqueous media and living cells. This work not only reported a robust molecule, RC, to disentangle the intricate interplay networks among different biothiols in biosystems but also demonstrated a strategy for the design of multisignal fluorescent probes for the simultaneous sensing of diverse RSS species as well as other biological species.