A ratiometric two-photon fluorescent probe for selective detection of endogenous formaldehyde via Aza-Cope reaction

An Aza-Cope reaction-based ratiometric two-photon fluorescent probe, MQAP, for formaldehyde detection was developed. [Display omitted] •MQAP showed selective ratiometric detection signal for formaldehyde.•MQAP has deep tissue penetration and low viability cytotoxicity.•MQAP exhibited a large two-photon absorption cross section at 720 nm.•MQAP can monitor endogenous formaldehyde in living cells and living zebrafish. A ratiometric two-photon fluorescent probe, MQAP, based on 6-substituted quinoline skeleton, has been developed for the detection of formaldehyde (FA) in aqueous solution, living cells, and living zebrafish. After treatment with FA, the fluorescence emission wavelength of MQAP red-shifts significantly from 405 nm to 490 nm, and the emission intensity ratio (I490nm/I405nm) dramatically increases from 0.34 to 7.52 (about 22 times). The response mechanism was proposed to be FA-induced 2-aza-Cope rearrangement on MQAP, as confirmed by LC–MS, theoretical calculation, UV-vis and fluorescence spectra. MQAP is almost insensitive to the change of pH value within the biologically relevant pH range, with a large two-photon absorption cross-section at 720 nm (185 GM). Additionally, MQAP has increased tissue penetration depth up to 150 μm, and low cytotoxicity to living cells. To demonstrate its biological applicability, MQAP was successfully used to detect endogenous FA in living cells and living 5-day-old zebrafish.