Synthesis And Research On Application Of Two-photon Fluorescent Sensors Based On Styrene

With the developments of technologies based on two-photon, two-photon fluorescence microscopy is becoming an essential imaging system in the research on life science. In comparison with one-photon confocal imaging, two-photon fluorescence microscopy can offer many advantages including NIR (near infrared) photons as excitation source, imaging in the black background, avoidance of photodamage and photobleaching, fixed target excitation, high transverse and longitudinal resolutions, small absorption coefficient of light in tissue, lower tissue autofluorescence, etc. As the two-photon (TP) absorption process is quadratically intensity-dependent, the excitation is confined to a small volume in the focal plane. Furthermore, the use of an infrared laser source improves light penetration depth in tissues, thus allowing deeper tissue imaging. Two-photon fluorescent probes applied to studies on the contents of ions and the effects of their contents upon physiology, physiological action mechanisms of ions, interplays between ions and molecules, distributions of specific molecules and their interactions, and so on, play an important role in imaging. Therefore, Two-photon fluorescent probes with superior photophysical properties are still required for imaging of biomedical field.The stilbene TP Zn2+ratiometric fluorescent probes BZn and DZn based on ICT mechanism have been designed and synthesized. The formation of complex BZn-Zn and DZn-Zn weakened the electron-donating character, leading to a blue shift in absorption and fluorescence emission. The TP action spectra of BZN-Zn indicated a 18-fold larger than those of TSQ-Zn. It can also be used in two-photon ratiometric fluorescence microscopy in living HEK293 cells. DZn is selective for over biologically competing alkali-and alkaline-earth-metal ions, and can distinguish Zn2+ from Cd2+ by naked-eye sand fluorimetric experiments. The TP action spectra of DZn-Zn indicated aΦδvalue of ca.185GM at 830 nm,40-fold larger than those of TSQ-Zn. DZn was confirmed to be directly applicable to monitor changes in intracellular Zn2+ in living tissues, owing to its membrane permeability. TP ratiometric imaging with DZN at a depth of 120μm could clearly visualize endogenous Zn2+ in hippocampal slices without suffering from dye localization artifacts.New water-soluble TP ratiometric fluorescent probe DCN4 for Pb2+ containing a 1,4-dicyano-2,5-bis(styryl) benzene fluorophore and 2-(N,N'-bis(carboxylmethyl)) amino-1-carboxylmethoxyl benzene as receptor has been synthesized. With the addition of different concentrations of lead ion, the broad emission intensities at 590 nm gradually increase, and TP action cross section (Φδ) is 51 GM at 740 nm. Novel TP fluorescent probe DCN2 based on hydrophobic effect mechanism has been designed and synthesized to detect HSA (human serum albumin). The formation of complex DCN2-HSA leading to a blue shift of 73 nm in fluorescence emission and an increase of quantum yield for about 90 fold. A good linear relationship was observed up to 20μM(1320μg ml-1) of protein. The staining procedure of DCN2 required only 30 min and fluorescent image of proteins was observed despite the removing of SDS and excess dyes in the gel. The TP action spectra of DCN2-HSA indicated aΦδvalue of ca.85GM at 830 nm TPM (two-photon microscopy) images were obtained from living cells and tissues. Moreover, by using TPM, this probe is capable of monitoring proteins at a depth of about 180μm in living tissues.