A Nile Blue based near-infrared fluorescent dye (RB-S) was reported, which was synthesized by a simple one-step reaction between 5-(dimethylamino)-2-nitrosophenol and 1-naphthaldehyde under mild reaction conditions. RB-S was well characterized using 1H NMR, 13C NMR, and TOF-MS. RB-S showed a significative fluorescent response to different concentrations of homocysteine or cysteine in the acetonitrile-water solution (4:1, V/V, r.t.). Furthermore, with the concentration of homocysteine or cysteine increasing, the fluorescent intensity of RB-S at 755 nm increased gradually, and that at 685 nm decreased. Exhilaratingly, there was a linear ratio fluorescent response (I755 nm/I685 nm) to the concentration of homocysteine or cysteine in the range of 0.03～0.33 μmol/L. The recognition mechanism of the ratio fluorescent response to homocysteine or cysteine was verified by two experiment methods (high performance liquid chromatography-mass spectrum hyphenation technology and 1H NMR titration) in this paper. A lot of detailed experiment results indicated that the ratio fluorescent response of RB-S to homocysteine or cysteine is most likely the result of the cyclization reaction of the aldehyde group to a thiazolidine group. The detection limit of RB-S for homocysteine or cysteine was determined as 0.025 μmol/L (3σ/k) in the acetonitrile-water solution (4:1, V/V, r.t.). Besides sensitivity, the selectivity of RB-S for homocysteine or cysteine was another very important parameter to evaluate in this paper. The selectivity experiments of RB-S were extended to 51 kinds of other biological-related species, including ion, amino acid and so on. All the experiment results demonstrated that RB-S was highly selective for homocysteine or cysteine over other related species and could meet the selective requirements for the fluorescent bioimaging and detecting of homocysteine or cysteine in vivo and in vitro. Therefore, RB-S was successfully used to ratio fluorescent detect and image homocysteine or cysteine in serum samples and living cells by fluorescence spectrum and confocal fluorescence microscopic imaging, respectively.
王魁, 刘自力, 蒋凯. 比率检测高/半胱氨酸荧光染料的合成及活细胞成像研究[J]. 化学学报, 2014, 72(5): 590-594.
Wang Kui, Liu Zili, Jiang Kai. Synthesis and Imaging Performance Research of a Ratio Fluorescent Dye for the Determination of Homocysteine/Cysteine in Living Cell. Acta Chimica Sinica, 2014, 72(5): 590-594.