The Construction Of Near-infrared Fluorescent Probes Based On Hemicyanine And Their Bioimaging Applications In Reactive Sulfur Species

Hemicyanine dyes,with highly diverse structural tenability,are promising fluorophores for near-infrared（NIR）bioimaging,can be modified to reach high fluorescence quantum yield and good biocompatibility which is suitable for bioimaging.Recently,a growing number of studies have investigated the development of hemicyanine-based NIR probes in response to various biomarkers.Intracellular redox homeostasis provides implications in physiological and pathological fields,the disruption of redox homeostasis is closely associated with some human diseases.Many reactive sulfur species（RSS）play important roles in maintaining redox homeostasis,serving as antioxidants and acting as free radical scavengers.Cys is the only amino acid that is involved in enzyme active sites,protein and peptide construction.It is also an important biomolecule in regulation of biological nitrogen balance and redox homeostasis.Detection of Cys concentration has clinical implications as a biomarker for several diseases.H₂S,an essential gas signal molecule,has been reported to function in many physiological processes,including regulating inflammation,relaxation of vascular smooth muscles,mediation of neurotransmission,and redox status modulation.Although H₂S as a cell protectant and gas transport agent is involved in a variety of diseases and has multiple functions,its role and mechanism of action are still unclear.Therefore,it is helpful to promote disease prevention and clinical diagnostics by developing effective molecular tools to monitor intracellular RSS in real time and investigate response mechanisms.In this thesis,we have designed and synthesized fluorescent probes based on hemicyanine for the detection of Cys and H₂S,and explore their imaging applications in cells.The research works were as follows:（1）We reported a cascading-response fluorescent probe based on hemicyanine,Cy-Cys-p H,for the sequential detection of Cys and p H in pancreatic cancer cells.In the presence of Cys,Cys-mediated cleavage of the acrylate group caused Cy-Cys-p H to be transformed into Cy-Cys-O,which induced intense fluorescence enhancement at 725 nm.Then,Cy-Cys-O was protonated to obtain Cy-Cys-OH and the fluorescence emission shifted to 682 nm,showing a ratiometric p H response.Furthermore,Cy-Cys-p H can monitor the intracellular p H during the therapeutic process with anticancer drugs and evaluated the ability of three anticancer drugs to kill Panc-1 cells,proving that associating Cys and p H is in part an effective anticancer strategy in the treatment of pancreatic cancer.Significantly,Cy-Cys-p H is able to monitor and image p H changes during Cys depletion in real-time,which further reveals the molecular mechanism of Cys-depleted pancreatic cancer cell death,providing a powerful molecular tool for the precise treatment of PC.（2）We reported a ratiometric fluorescent probe based on hemicyanine,Cy-H₂S,for detecting H₂S rapidly in cells.We connected the aldehyde group to the ortho-position of 2,4-dinitrophenyl ether,which is the response site of H₂S.The fast nucleophilic addition of H₂S to an aldehyde group was combined with the subsequent intramolecular thiolysis of dinitrophenyl ether in order to be used to detect H₂S efficiently.Cy-H₂S with simple structure and high sensitivity,it responded to H₂S in only 150 seconds,which was 16 times faster than the probe in ortho-position of 2,4-dinitrophenyl ether without the aldehyde group.Meanwhile,Cy-H₂S has the potential to be used for quantitative detection of H₂S because of its colorimetric and ratiometric properties.Notably,Cy-H₂S successfully imaged intracellular H₂S changes induced by the enzymes,evaluated the release efficiency of the H₂S prodrug,and revealed the increase and inhibition of NO-associated H₂S.It provides a new and effective tool for the study of H₂S prodrugs in vivo and the sensing mechanism between gas signal molecules.