Design,Synthesis And Bioactivity Study Of Novel Fluorescent Probes Based On Dihydroxanthene Skeleton

Fluorescence analysis technology is one of the important tools of detecting and imaging analytes,which has been given rise to growing interest in the field of chemical biology and medicine in recent years ascribe to the advantages of convenient operation,high selectivity,high sensitivity,and dynamic monitoring in real time.This technology has attracted more and more attention for investigating and revealing the regulation mechanisms of related active substances on various physiology and pathology at the molecular levels.As one of the most important carriers of fluorescence analysis technology,the fluorescence performances of probes determine their application prospect.At present,fluorescent probes with long-emission wavelengths have attracted great interest in the development and application of such fluorescent probes because of their low interference of background fluorescence signal,strong tissue penetration ability and little damage to cells.Dihydroxyanthracene,as a derivative of oxyanthracene,has long conjugated structures that can be modified and multiple structural sites that can regulate the fluorescence properties of probes.Therefore,fluorescent probes based on dihydroxyanthracene design exhibit outstanding advantages in the detection and imaging of many active substances.Although the small molecule probes containing the dihydroxyanthracene skeleton have been reported in the literatures,these probes still show some obvious shortcomings in practical applications,for example,small Stokes shift is to lead to self-absorption that is not conducive to bioimaging analysis.Therefore,it is still necessary to take the advantages of dihydroxyanthracene skeleton to reasonably design and synthesize new fluorescent probe molecules in order to enrich the probe molecular library and expand the detection application of active molecules.In this paper,by modifying the dihydroxyanthracene skeleton reasonably,the following works were completed:1.By introducing functional pyridine salt as the receptor moeity into dihydroxyanthracene skeleton,a functional fluorescent probe DHX-1 with mitochondria-targeted was designed and synthesized,for the"turn off"near-infrared fluorescence response to hydrogen peroxide.The spectral changes of the probe response to hydrogen peroxide were measured by UV and fluorescence spectra,and the colorimetric and fluorescence quantitative detections of hydrogen peroxide were realized at the emission wavelength of 675 nm,and the detection limit of probe was obtained to be 4.21μM.Secondly,the response mechanism of the probe to hydrogen peroxide was investigated by UV spectroscopy analysis and DFT calculation.Based on its excellent optical properties,the probe successfully realized the imaging detection of endogenous and exogenous hydrogen peroxide in both of living cells and zebrafish.2.By introducing N,N-diethylamino group as donor and two different groups,namely aldehyde group and active thiazole vinyl group,as acceptors as well as recognition site into dihydroxyanthracene skeleton,two fluorescent probes DHX-2 and mitochondria-targetable DHX-3 based on intramolecular charge transfer mechanism were constructed respectively for the"on-off"near-infrared fluorescence response to SO₂ at the emission wavelength of650 nm and 748 nm.The results of spectral experiments showed that both types of probes could realize the selectively quantitive detection of SO₂,and the detection limits of probes are 5.6μM（DHX-2）and 4.08μM（DHX-3）respectively.Cells and zebrafishs experiments confirmed that the two types of probes have successfully realized the imaging detection of endogenous and exogenous SO₂ in organisms ascribe to their excellent near-infrared fluorescence performances.Moreover,DHX-3 also successfully realized the monitoring of SO₂ owing to its advantages of long wavelength emission in vivo.3.By coupling of phenothiazine with electron rich system as donor into dihydroxyanthracene,a novel fluorescent probe DHX-4 with PET effect was constructed for"turn on"response to hypochlorous acid.The phenothiazine moiety of probe DHX-4 not only tunes the fluorescence performances of probe,but also acts as a specific recognition site of hypochlorite,resulting in changes of its UV and fluorescence spectras that realizes the colori/fluorimetric detection of hypochlorous acid at the emission wavelength of 600nm.Thus,the detection limit of probe is 2.33μM for hypochlorite.In addition,probe DHX-4 also successfully realized the imaging detection of endogenous and exogenous hypochlorous acid in cells and zebrafish,showing excellent imaging performance.