Chemiluminescent Probes for Activity‐Based Sensing of Formaldehyde Released from Folate Degradation in Living Mice

Formaldehyde (FA) is a common environmental toxin that is also produced naturally in the body through a wide range of metabolic and epigenetic processes, motivating the development of new technologies to monitor this reactive carbonyl species (RCS) in living systems. Herein, we report a pair of first‐generation chemiluminescent probes for selective formaldehyde detection. Caging phenoxy‐dioxetane scaffolds bearing different electron‐withdrawing groups with a general 2‐aza‐Cope reactive formaldehyde trigger provides chemiluminescent formaldehyde probes 540 and 700 (CFAP540 and CFAP700) for visible and near‐IR detection of FA in living cells and mice, respectively. In particular, CFAP700 is capable of visualizing FA release derived from endogenous folate metabolism, providing a starting point for the use of CFAPs and related chemical tools to probe FA physiology and pathology, as well as for the development of a broader palette of chemiluminescent activity‐based sensing (ABS) probes that can be employed from in vitro biochemical to cell to animal models. Chemilumineszierende aktivitätsbasierte Sonden der ersten Generation für die In‐vivo‐Bildgebung von Formaldehyd (FA) wurden entwickelt. Ein FA‐selektiver Auslöser wird mit einem Phenol‐Dioxetan‐Gerüst kombiniert, das nach einer Aza‐Cope‐abhängigen Aktivierung ein Photon emittiert. Mit diesen Reagentien ließen sich FA‐Flüsse in lebenden Zellen und Mäusen darstellen, was die Identifizierung einer endogenen FA‐Produktion aus metabolischen Folatzyklen in vivo ermöglicht.