Design,Synthesis And Application Of Organic Small Molecule Fluorescent Probes For Cysteine?Cys? And Sulfur Dioxide?SO₂?

As a kind of fluorescent probes,organic small molecule fluorescent probes have become important tools for small molecule detection in biological samples due to excellent selectivity,high sensitivity,rapid bio-permeability and real-time imaging.Selecting known fluorophores or constructing new ones and using known or innovative mechanisms to build organic small molecule fluorescent probes are currently the main strategies in this direction.Cysteine(Cys)and sulfur dioxide(SO2)are two important small molecules in the metabolism of Methionine(Met).Moreover,SO2 is a metabolite of Cys.Cys is a conditionally essential amino acid and glycogenic amino acid in human body.It is also a synthetic precursor of various substances in living body and can reflect the degree of oxidative damage of the organism.Abnormal content of Cys may lead to diseases such as rheumatoid arthritis,Parkinson's disease,Alzheimer's disease,edema,growth retardation,liver damage and so on.SO2 is not only an important atmospheric pollutant,but also considered to be another important gas signal molecule after hydrogen sulfide(H2S),carbon monoxide(CO)and nitrogen monoxidum(NO).It maintains the redox balance and sulfur balance in the body,regulates insulin levels in the blood and reduces blood pressure.Furthermore,excessive inhalation of SO2 can also cause a variety of diseases,such as lung cancer,myocardial ischemia,stroke,brain cancer and so on.Therefore,the development of organic small molecule fluorescent probes for the measurement of Cys and SO2 levels in organisms is of great benefit for physiological function assessment and related disease warning.Based on the current research hotspots,literature research,laboratory research foundation and my own interests,we constructed two new organic small molecule fluorescent probes:QP-1 and NJUXJ-1,for the detection of Cys and SO2,respectively.The fluorophore of QP-1 was a quinoxalinone derivative which was used as a fluorophore for the first time and the recognition mechanism was the specific addition cyclization reaction between Cys and acrylate.On the other hand,the fluorophore of NJUXJ-1was chosen as N,N-dimethylquinolin-6-amine based on laboratory study and the recognition mechanism was the specific Michael addition reaction between SO2 and carbon-carbon double bond.By measuring the absorption spectrum,emission spectrum,reaction time,pH stability,sensitivity,selectivity,interference and fluorescence quantum yield of QP-1 and NJUXJ-1,we conducted a comprehensive in vitro property evaluation.Based on these results,we carried on the preliminary safety assessment and further performed application assessment.After QP-1reacted with Cys,the fluorescence spectrum underwent a red shift of 70 nm;it effectively eliminated the interference of other small molecule biothiols(Hcy,GSH),realized the qualitative identification of exogenous and endogenous Cys in HeLa cells and was successfully applied in rat urine model;the biggest innovation point of QP-1 was its new fluorophore,which was helpful for the development of organic small molecule fluorescent probes with unique fluorescence properties and further improvement of the probe practicability.As for NJUXJ-1,after reacting with S02,the fluorescence spectrum underwent a blue shift of 80nm and the fluorescent intensity increased by more than 60 times;it had an excellent detection limit(13 nM)and realized the quantitative detection of exogenous SO2 and qualitative detection of endogenous SO2 in HeLa cells;the most attractive point was the rapid cell permeability(2min)of NJUXJ-1,which greatly shortened the incubation time required for cell imaging,effectively reduced background interference and provided a more convenient way for deeper applications.The development of the two tools provided potential methods for systematic investigation of sulfur metabolism in living organism,which is of great value for the evaluation of related fields.