A, Wrobel.; T, Johnstone.; A, Liang.; S, Lippard.; Pablo Rivera-Fuentes. J. Am. Chem. Soc. 2014, 136, 4697
DOI: 10.1021/ja500315x
The first near-infrared fluorescent turn-on sensor for the detection of nitroxyl (HNO), the one-electron reduced form of nitric oxide (NO), is reported. The new copper-based probe, CuDHX1, contains a dihydroxanthene (DHX) fluorophore and a cyclam derivative as a Cu(II) binding site. Upon reaction with HNO, CuDHX1 displays a five-fold fluorescence turn-on in cuvettes and is selective for HNO over thiols and reactive nitrogen and oxygen species. CuDHX1 can detect exogenously applied HNO in live mammalian cells and in conjunction with the zinc-specific, green-fluorescent sensor ZP1 can perform multicolor/multianalyte microscopic imaging. These studies reveal that HNO treatment elicits an increase in the concentration of intracellular mobile zinc.
Introduction
HNO molecule not only play very important role in living organism but also is excretion molecule that results from variety of biological activities. Therefore, to visualize their pinpoint localization enables researcher to have a deeper understanding of Endogenous bioactivity. Although some dyes that selectively detect the presence of NO is now available from chemical companies, the advent of new near-infrared emission dye with high selectivity is highly is anticipated.
Working mechanism
Incorporating Cu atom with nitrogen chelating agent is widely used strategy to prepare HNO fluorescent sensitizer like CuBOT1 and CuCOT1.(1)(2) In the chelated state, the photo-induced electron transfer (PET) from the dye’s singlet excited state to the bound Cu2+ ion occurs, resulting the fluorescence OFF-state. Once the Cu2+ are reduced to Cu+ by NO molecule, the Cu atoms take apart and leave the dyes in fluorescence OFF-state. By applying same strategy, the group achieved the first near-infrared fluorescent turn-on sensor for the detection of HNO as shown below
Molecular design
(1) Commercial available IR-780 was used as starting material.
(2) Target compound DHX1 with long conjugated system (highlighted in red) was obtained.
(3) Cyclam (1,4,8,11-tetraazacyclotetradecane) was chosen as copper chelate side.(3)
Selectivity and spectrum
Various oxidant reagents including were tested for CuDHX1 to see the selectivity against HNO. As can be seen below, CuDHX1 was the most sensitive to HNO. The increase in fluorescence intensity and corresponding physical data are summarized below.
Multicolor imaging
To confirm its adaptability and functionality for practical uses, CuDHX1 (red emission) in combination with Hoechst 33528 dye (Blue DNA stainer) and ZP1 (green zinc sensitizer) was used to stain Hela cell. Addition of appropriate reagents for fluorescence activation revealed that CuDHX1 worked without any interference in other dyes. As shown in the last picture, three dyes functioned independently and could be detected through each channel.
Reference
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Rosenthal, J.; Lippard, S. J. J. Am. Chem. Soc. 2010, 132, 5536. DOI: 10.1021/ja909148v
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