Mechanistic Investigation on Chemiluminescent Formaldehyde Probes.

A first-generation pair of chemiluminescent formaldehyde (FA) probes (CFAP540 and CFAP700) was reported recently. CFAP540 and CFAP700 with high selectivity and sensitivity to FA, are respectively suitable in cell and in vivo. Experimentalists have confirmed that both probes utilize a general 2-aza-Cope FA-reactive trigger and a chemiluminogenic phenoxydioxetane scaffold. The mechanism and detailed process of CFAP chemiluminescence (CL) remain largely unknown. In the present paper, we performed (time-dependent) density functional theory calculations on the entire reaction process of CFAP540 with FA to produce CL. The calculations elucidated the CL producing process: FA initiates the decomposition of CFAP540 by dehydration condensation, and a phenoxy 1,2-dioxetane is formed through a series of reactions of aza-Cope rearrangement, hydrolysis of imine, and β-elimination of alkoxyl group. Afterwards, the produced phenoxy 1,2-dioxetane decomposes to produce m-oxybenzoate derivative at the first singlet state (S1) via two crossings between potential energy surfaces of the ground state (S0) and S1 state. This m-oxybenzoate derivative was assigned to the light emitter of the CFAP540 CL. The CL producing process and assignment of the light emitter of CFAP700 CL are similar with the corresponding ones of CFAP540. By analyzing the D-π-A architecture of the light emitters of CFAP540 and CFAP700, we theoretically designed a series of CFAPs and proposed a scheme to modulate their CL from visible to near-infrared region by adjusting the length and structure of the π-bridge.