Study On Synthesis And Properties Of Two-photon Hydrogen Sulfide Fluorescent Probe

Hydrogen sulfide (H2S) has emerged as the third endogenous gaseous signaling compound, which is associated with diverse physiological and pathological processes. The production and metabolism of H2S in mammalian cells are regulated by several enzymes such as cystathionine ?-lyase (CSE), cystathionine ?-synthase (CBS), cysteine aminotransferase (CAT) and 3-mercaptopyruvate sulfurtransferase (3-MST). Once these enzymatic reaction imbalances will cause the abnormal level of H2S, leading to various diseases, like Down's syndrome, Alzheimer's disease. The traditional technology of H2S detection includes methylene blue method, electrochemical and chromatograpgy methods. However, these methods require the living bio-samples to be post-mortem processed and destructed. By contrast, emerging fluorescence detection technology has incomparable advantages in comparison to the traditional detecting technology. Therefore, it is particularly important to develop a fast response, high sensitivity/selectivity, non-invasiveness, and aptness for living cells, tissues.Benzimidazoles are very useful building blocks for the fluorescent probe that is applied to detect biological molecules or ions, in addition to the application of the medicine. 2-(2-hydroxyphenyl)benzimidazole is known to be excited state intramolecular proton transfer (ESIPT), existing a ketone and enol tautomerism. In addition, it has large Stokes shift, high fluorescence quantum yield and can be used for two-photon microscope imaging. However, the fluorescence probe based on benzimidazoles is less reported. In this paper, we choose the 2-(2-hydroxyphenyl)benzimidazole as the fluorophore, piperazine derivatives are introduced to the 5-site. Taking advantage of the strong nucleophilic property, we design and synthesize two novel two-photon H2S fluorescence probes basing on the dinitrophenyl ether thiolysis.5-Bromo-2-nitroacetanilide was synthesized by nitration of m-bromoaniline. It reacted with 4-methylpiperazine/4-(2-hydroxyethyl)piperazine and hydrolysis to give 5-(4-methyl piperazinyl)-2-nitroaniline and 5-(4-(2-hydroxyethyl)piperazinyl)-2-nitroaniline, respectively. They were converted into o-phenylenediamines through hydrogenation catalyzed by Pd/C. The o-phenylenediamines reacted with salicylaldehyde to afford 5-(4-methylpiperazinyl)-2-(2-hydroxylphenyl)benzimidazole and 5-(4-(2-hydroxyethyl)piperazinyl)-2-(2-hydroxyl phenyl)benzimidazole, respectively. Finally the two compounds reacted with 2,4-dinitro-bromobenzene to generate two target compounds,5-(4-methylpiperazinyl)-2-(2-(2,4-dinitro phenoxy)phenyl)benzimidazole (compound 1) and 5-(4-hydroxyethylpiperazinyl)-2-(2-(2,4-dinitrophenoxy)phenyl)benzimidazole (compound 2), respectively. The properties of the target compounds were studied through testing its optical properties, fluorescence titration, ion selective, fluorescence imaging and cell cytotoxicity. The determinations indicated that above two probes had high sensitivity (2.2 ?M,2.3?M, respectively), high selectivity (in test species, only the H2S has obvious response), quicker response speed (30 min) and good bio compatibility (under test concentration, the cell viability was as high as 90%, and even when the concentration reached 160 mol/L, the cell activity was still more than 70%). Compound 2 had larger background interference and a slower response speed.