Despite the promising photofunctionalities, phosphorescent probes have been examined only to a limited extent, and the molecular features that provide convenient handles for controlling the phosphorescence response have yet to be identified. We synthesized a series of phosphorescence zinc sensors based on a cyclometalated heteroleptic Ir(III) complex. The sensor construct includes two anionic cyclometalating ligands and a neutral diimine ligand that tethers a di(2-picolyl)amine (DPA) zinc receptor. A series of cyclometalating ligands with a range of electron densities and band gap energies were used to create phosphorescence sensors. The sensor series was characterized by variable-temperature steady-state and transient photoluminescence spectroscopy studies, electrochemical measurements, and quantum chemical calculations based on time-dependent density functional theory. The studies demonstrated that the suppression of nonradiative photoinduced electron transfer (PeT) from DPA to the photoexcited Ir(IV) species provided the underlying mechanism that governed the phosphorescent response to zinc ions. Importantly, the Coulombic barrier, which was located on either the cyclometalating ligand or the diimine ligand, negligibly influenced the PeT process. Phosphorescence modulation by PeT strictly obeyed the Rehm-Weller principle, and the process occurred in the Marcus-normal region. These findings provide important guidelines for improving sensing performance; an efficient phosphorescence sensor should include a cyclometalating ligand with a wide band gap energy and a deep oxidation potential. Finally, the actions of the sensor were demonstrated by visualizing the intracellular zinc ion distribution in HeLa cells using a confocal laser scanning microscope and a photoluminescence lifetime imaging microscope.

Phosphorescence signaling provides a valuable alternative to conventional bioimaging based on fluorescence. The benefits of using phosphorescent molecules include improved sensitivity and capabilities for effective elimination of background signals by time-gated acquisition. Cyclometalated Ir(III) complexes are promising candidates for facilitating phosphorescent bioimaging because they provide synthetic versatility and excellent phosphorescence properties. In this Forum Article, we present our recent studies on the development of phosphorescence sensors for the detection of metal ions based on cyclometalated iridium(III) complexes. The constructs contained cyclometalating (C^N) ligands with the electron densities and band-gap energies of the C^N ligand structures systematically varied. Receptors that chelated zinc, cupric, and chromium ions were tethered to the ligands to create phosphorescence sensors. The alterations in the C^N ligand structures had a profound influence on the phosphorescence responses to metal ions. Mechanistic studies suggested that the phosphorescence responses could be explained on the basis of the modulation of photoinduced electron transfer (PeT) from the receptor to the photoexcited iridium species. The PeT behaviors strictly adhered to the Rehm-Weller principle, and the occurrence of PeT was located in the Marcus-normal region. It is thus anticipated that improved responses will be obtainable by increasing the excited-state reduction potential of the iridium(III) complexes. Femtosecond transient absorption experiments provided evidence for the presence of an additional photophysical mechanism that involved metal-ion-induced alteration of the intraligand charge-transfer (ILCT) transition state. Utility of the mechanism by PeT and ILCT has been demonstrated for the phosphorescence sensing of biologically important transition-metal ions. In particular, the phosphorescence zinc sensor could report the presence of intracellular zinc pools by using confocal laser scanning microscopy and photoluminescence lifetime imaging microscopy techniques. We hope that the significant knowledge gained from our studies will be of great help in the design of new molecules as phosphorescence sensors.

We have developed a series of molecular probes for the fluorescent detection of singlet dioxygen ((1)O2). The probes, based on asymmetrically substituted 1,3-diarylisobenzofurans, undergo the [2 + 4] cycloaddition reaction with (1)O2, producing ratiometric fluorescent responses. Two-photon fluorescence microscope experiments demonstrated the biological utility of the probes for the visualization of endogenous (1)O2 in macrophage cells.

A new fluorescence turn-on probe that responds exclusively to H(2)O(2) exhibits subcellular localized fluorescence staining of lysosomes.