Novel Hydrogen Sulfide Donor And Fluorescent Probe For?-Glutamyl Transpeptidase Detection

Hydrogen sulfide molecule has been identified as a cell-signaling mediator, which was involved in various physiological and phthological processes. It is essential to maintain Hydrogen sulfide concentration in balance in vivo. All of these findings suggest that regulation of endogenous H2S formation and exogenous H2S administration may have therapeutic benefits. In this field, H2S donors (also known as H2S releasing agents) are important tools. GGT is a cell membrane-bound enzyme, which selectively catalyzes the cleavage of the y-glutamyl bond in GSH, resulting in the production of cysteinyl-glycine, followed by formation of cysteine and glycine induced by plasma membrane dipeptidase. It has been discovered that overexpressed levels of GGT are associated with tumorigenesis in several human cancer cells, as represent by cervical and ovarian cancers.Thus, GGT can be regarded as a biomarker for cancer diagnosis as well as a therapeutic target.This work mainly covers the following three parts:1. Novel light-activated hydrogen sulfide donor releasing out hydrogen sulfide and coumarin fluorophore, was prepared from nitrobenzyl mercaptan, 4-hydroxymethyl-7-methoxy-coumarin and levulinic acid, and its structure was characterized by 1H NMR,13C NMR and MS. The H2S donor freed non-hydrogen sulfide and non-fluorescence. After irradiated by the 365 nm UV light irradiation, the H2S donor liberated hydrogen sulfide rapidly and reached a peak, and emitted remarkable fluorescence while the H2S donor was excited by 365 nm UV light. When the H2S donor was under the irradiation by the 365 nm UV light for 3-9 min, the fluorescence intensity and the concentration of hydrogen sulfide was dynamics of correlation. By detecting changes in fluorescence intensity, the release of hydrogen sulfide can be monitored indirectly.2. An NBD-Cl-based fluorescent probe for GGT was prepared from NBD-Cl and glutathione, and its structure was characterized. With the 475 nm UV light irradiation excitation, the probe showed faint fluorescence. After the probe reacted with GGT, it glows bright green fluorescence and the maximum emission wavelength was 538 nm, with the same excitation wavelength (475 nm). The probe has a high stability, high selectivity at physiological pH conditions. The fluorescence intensity is dependent on the concentration of GGT with the linear response range being 1-40U, the detection limit of GGT is 0.13U/L. Confocal imaging experiments show that the probe can be used for visual inspection of living cells GGT.