Ligand-Directed Approach to Activity-Based Sensing: Developing Palladacycle Fluorescent Probes That Enable Endogenous Carbon Monoxide Detection

Carbon monoxide (CO) is an emerging gasotransmitter and reactive carbon species with broad anti-inflammatory, cytoprotective, and neurotransmitter functions along with therapeutic potential for the treatment of cardiovascular diseases. The study of CO chemistry in biology and medicine relative to other prominent gasotransmitters such as NO and H₂S remains challenging, in large part due to limitations in available tools for the direct visualization of this transient and freely diffusing small molecule in complex living systems. Here we report a ligand-directed activity-based sensing (ABS) approach to CO detection through palladium-mediated carbonylation chemistry. Specifically, the design and synthesis of a series of ABS probes with systematic alterations in the palladium-ligand environment (e.g., sp³-<i>S</i>, sp³-<i>N</i>, sp²-<i>N</i>) establish structure-activity relationships for palladacycles to confer selective reactivity with CO under physiological conditions. These fundamental studies led to the development of an optimized probe, termed Carbon Monoxide Probe-3 Ester Pyridine (COP-3E-Py), which enables imaging of CO release in live cell and brain settings, including monitoring of endogenous CO production that triggers presynaptic dopamine release in fly brains. This work provides a unique tool for studying CO in living systems and establishes the utility of a synthetic methods approach to activity-based sensing using principles of organometallic chemistry.