Design Of Near-infrared Fluorescent Probe Based On Cyanine And The Applications For Imaging

As an efficient technique suitable for visualization in vivo, fluorescence imaging applied to monitor small molecules in biological samples. The near-infrared fluorescence imaging based on organic small molecular probes has shown extensive and attractive application prospect on account of low background, deeper penetration of tissue, relatively high sensitivity, and less organism photodamage. The work of this dissertation focused on design and synthesis of near –infrared fluorescence sensors based on cyanine skeleton for quantitative determination of mercapto amino acids（GSH, Cys, and Hcy）, N2H4, and O₂^·-.The main works are as follows:（1） The near-infrared（NIR） dyes detecting intracellular GSH would be helpful tools to understand the mechanisms of diseases. In chapter two, We report a facile method to design cyanine-based fluorescent probe in terms of biothiol-promoted specific O-S cleavage of the probe CyDNS and then self-immolate through an intramolecular 1,6-elimination. In addition, we have successfully applied the probe to living cells, in vivo and various tissues.（2） We report a specific probe with turn-on fluorescence property, visible color change with naked-eye and large wavelength shift on UV spectra for highly selective detection of Cys over homocysteine（Hcy） and glutathione（GSH） both in HEPES buffer（10 mM, pH 7.4） and diluted human serum. The probe that based on the conjugate addition-cyclization reaction has a low limit of detection to Cys（0.16 μM as NIR fluorescence sensor and 0.13μM as UV sensor）. Kinetic study indicated that the probe has a very rapid response to Cys owing to the much higher pseudo-first-order reaction constant with Cys（299 M-1s-1） than with Hcy（1.29 M-1s-1） or GSH（0.53 M-1s-1）. Upon addition of Cys to a solution of the probe, it happened a color change from purple to cyan, the maximum wavelength shifting from 582 nm to 674 nm on UV spectrum and a fluorescence emission at 697 nm appear. It has been successfully applied for determination of Cys in diluted serum and bioimaging of Cys in living cells with low cell toxicity.（3） The near-infrared（NIR） fluorescence probe CyJ with a turn-on fluorescence based on the structure–emission property relationships of the NIR dyes containing an acetyl group as recognizing moiety to N2H4. This new probe not only can be readily prepared, but also shows excellent sensing properties. First and most important of all, CyJ is highly selective for N2H4 over various anions, cations and other amino compounds and has a low limit of detection（LOD） of hydrazine（5.4 ppb as fluorescence sensor and 6.1 ppb as UV sensor）. Besides, CyJ exhibited a dramatic increase in fluorescence at λmax = 706 nm in the presence of N2H4, and it offers a rapid, colorimetric and vapor sensing detection process for N2H4 in both aqueous solution and diluted human serum. Furthermore, CyJ has good cell-membrane permeability and low cytotoxicity. In addition, we have successfully applied the CyJ to visualize N2H4 in live mouse and, for the first time, in tissues such as the liver, lung, kidney, heart, and spleen.（4） We developed a near-infrared（NIR）, turn on fluorescence probe CyR containing a phosphinate group as recognizing moiety for detecting O₂^·- , which took advantage of the nucleophilic properties of O₂^·-. This new probe can be readily prepared, and shows eminent sensing properties. Also, CyR is highly selective for O₂^·- over other reactive oxygen species（ROSs） and reductants, with a low limit of detection（LOD, 9.9 nM）. Kinetic study illustrated that CyR has a rapid response to O₂^·- for only 10 min. Besides, CyR has good cell-membrane permeability, intracellular stability, and low cytotoxicity. In addition, we have successfully applied the CyR to visualize O₂^·- in live zebrafish, mouse and, for the first time, in mouse liver.