Study On Novel PH Fluorescent Probes And Their Application In Cells Imaging

Chapter 1,this chapter reviewed the functions and monitoring significance of intracellular pH?pHi?,methods to determine intraceular pH?pHi?and to deliver the fluorescent probes into cells,the typical sensing mechanism of small-molecule fluorescent probes and research progress of small-molecule fluorescent probes.Chapter 2,by Knoevenagel reaction,quinoline and benzothiazole were introduced as recognition group and fluorophore,respectively,forming the pH probe BTVQ based on ICT.Its molecular structure was demonstrated by NMR and mass spectrometry.Its optical properties were studied by the ultraviolet-visible absorption spectrometry and fluorescence spectrometry.Its absorption and fluorescence properties indicated that the probe displays an obvious emission at 428 nm??ex=295 nm?,but gradually quenches with the decrease in pH values from 7 to 2.2 without emission shift.FMIP displays a large Stokes shift?133 nm?,which can reduce the excitation interference.Moveover,nonlinear fit of the sigmoidal curve?emission intensity versus pH value?affords the probe a pKa of 3.52 and the probe responded linearly and rapidly to minor pH fluctuations within the range of 3.0-3.8.Thus,this probe could be potentially suitable for monitoring pH variations in the acidic environments in cell.The presence of metal cations such as Na+,K+,Ca²⁺,Mg²⁺ and other common transition metal cations didn't interfere with its fluorescent pH response.The fluorescent images of Siha cells indicated that BTVQ had fine biocompatibility and could visualize acidic pH fluctuations in live cells,providing a new analytical method for monitoring strong acidic range pH changes in live cells.Chapter 3,3-[3-?4-Fluorophenyl?-1-?1-methylethyl?-1 H-indol-2-yl]-?E?-2-propenal?FMIP?was commonly used as an important drug precursor.For structure concern,FMIP contains electron donating substituents?the nitrogen atom in indole?and electron-accepting substituents?-F?with the D-?-A structure that may be available to possess intramolecular charge transfer?ICT?fluorescent properties and the nitrogen atom of indole moiety is sensitive to protonation and deprotonation.In this chapter we developed its applications in terms of the pH fluorescent probe.The pH titration indicated that FMIP displayed ratiometric response to pH and as the pH decreases from 7.0 to 3.5,the emission ratio?I578 nm/I478 nm?also changes dramatically from 3.48 to 0.68.The probe with pKa 3.90 and linear response to strong-acidity range of 3.3-4.5 can detect pH changes in strong acid conditions.FMIP displays a large Stokes shift?163 nm at pH 7.0 and 113 nm at pH 3.5?,which can reduce the excitation interference.Application of FMIP to pH imaging in live cells indicating that the probe had good cell membrane permeability and could be used to visualize intracellular pH fluctuations with negligible autofluorescence.It broadens the types of extreme acidic intracellular pH fluorescent probes and the results provides us a strategy to construct ratiometric fluorescent probes based on ICT effect.Chapter 4,3-[3-?4-Fluorophenyl?-1-?1-methylethyl?-1 H-indol-2-yl]-?E?-2-propena?FMIP?as recognition group was bridged to benzothiazole by Knoevenagel reaction to build a new pH fluorescent probe FMTQ.The pH titration indicated that FMTQ displayed a remarkable emission enhancement at 526 nm when the pH was decreased from 7.0 to 2.2.FMTQ also displays a large Stokes shift of 116 nm,which can reduce the excitation interference.The sigmoidal fitting of the emission intensity at 526 nm versus pH value yields a pKa value of 3.5.Interestingly,there is a good linearity between emission intensity at 526 nm and pH in the range of 2.3-4.1,which is valuable for strong acid conditions study.FMTQ also shared some other desired properties,such as good selectivity and excellent photostability.The results suggested that FMTQ has potential application in detecting intracellular acidic pH quantitatively.