Synthesis And Application Of Organic Fluorescent Probes For The Detection Of Active Sulfur In Living Organisms

Fluorescent probes are a class of functional molecules that detect certain specific identifiers through changes in fluorescent signals.Due to its high sensitivity,high specificity,high spatial and temporal resolution and non-invasiveness,it has become a reliable detection tool and is widely used in the fields of food sample analysis,bioactive molecule detection and in vivo imaging.As an important class of reactive substances in living organisms,reactive sulfur species are involved in metabolic processes such as redox in living organisms,and many disease occurrences are related to the overexpression of reactive sulfur species.Therefore,it is of great importance to design and synthesize a class of fluorescent probes that can specifically recognize active sulfur species and monitor it visually.This thesis briefly describes the development of fluorescent probes,and summarizes the work of domestic and foreign groups on the detection of active sulfur,and designs and synthesizes two new fluorescent probes for the detection of active sulfur,and investigates their performance and applications.First of all,we constructed a dual-channel sensitive fluorescent probe TMN-NBD for the highly effective detection of biothiols.By rationally incorporating NBD-Cl into a dicyanoisophorone derivative(TMN-OH),TMN-NBD was designed and synthesized.Pure TMN-NBD shows no fluorescence due to the strong electron-withdrawing property of NBD.After incubating with Cys/Hcy,the fluorophore TMN-OH was released and a strong fluorescence at 663 nm in the red channel was observed.In the same instant,another product NBD-NR was also formed via substitution/ intramolecular rearrangement reaction and gave a green emission(556 nm)in the green channel.While for GSH,the fluorescence was only increased in the red channel,which belongs to the fluorophore TMN-OH,and the produced sulfur-substituted NBD(NBD-SR2)displayed a minimal green emission due to the relatively weak electron donating ability of sulfur-atom in its structure.In our later experiments,the probe exhibited good performance and was successfully used for tracking biothiols in living cancer cells and the zebrafish model.Inspired by the vitro experiments,cellular experiments were next performed and the probe showed very low toxicity to He La cells and could track the biothiols of cancer cells.Finally,the probe was successfully used in live imaging of zebrafish.Secondly,we designed and synthesized a dual-response fluorescent probe(ES-OH)for the rapid and sensitive detection of viscosity and sulfite.The probe uses indole salt as the electron-absorbing group and 5-(4-hydroxyphenyl)thiophene-2-carbaldehyde as the electron-donating group of the molecular rotor.Due to the presence of ICT effect in the D-π-A structure of the probe,thus showing a certain red fluorescence,when the probe responds with sulfite,C═C is attacked and undergoes an addition reaction,making the intramolecular conjugation blocked and the ICT effect disappears fluorescence off.In addition,when the environment is at low viscosity,the energy obtained by the probe is dissipated in the form of double bond rotation,and therefore exhibits a weak red fluorescence.In contrast,when the environment is at high viscosity,the rotation is blocked and the probe dissipates energy in the form of luminescence,so that a strong fluorescence can be observed.Based on this,we examined the potential application of the probe ES-OH in living organisms,and preliminary results showed that ES-OH has low cytotoxicity and good biocompatibility.In further experiments such as intracellular co-localization,the probe was found to have the effect of targeting and mitochondria,and to provide highly sensitive detection of the viscosity of subcellular organelle.It was further found in LPS induction experiments that the probe can provide highly sensitive feedback on viscosity changes during cellular inflammation.Also,the probe can respond rapidly to exogenous sulfite.Finally,we applied the probe to zebrafish in vivo for the same experiments and found that ES-OH also showed good results in vivo.In summary,we have successfully constructed two fluorescent probes for the detection of reactive sulphur,both of which have shown good performance and have been successfully applied in living organisms.Finally,it is hoped that they may provide a viable means of detection for clinical reactive sulphur-induced diseases.