Sydnthiones are versatile bioorthogonal hydrogen sulfide donors.

Hydrogen sulfide (HS) is an important endogenous gasotransmitter, but the bioorthogonal reaction triggered HS donors are still rare. Here we show one type of bioorthogonal HS donors, sydnthiones (1,2,3-oxadiazol-3-ium-5-thiolate derivatives), which was designed with the aid of density functional theory (DFT) calculations. The reactions between sydnthiones and strained alkynes provide a platform for controllable, tunable and mitochondria-targeted release of HS. We investigate the reactivity of sydnthiones‒dibenzoazacyclooctyne (DIBAC) reactions and their orthogonality with two other bioorthogonal cycloaddition pairs: tetrazine‒norbornene (Nor) and tetrazine‒monohydroxylated cyclooctyne (MOHO). By taking advantage of these mutually orthogonal reactions, we can realize selective labeling or drug release. Furthermore, we explore the role of HS, which is released from the sydnthione-DIBAC reaction, on doxorubicin-induced cytotoxicity. The results demonstrate that the viability of H9c2 cells can be significantly improved by pretreating with sydnthione 1b and DIBAC for 6 h prior to exposure to Dox.