Design And Synthesis Of Fluorescent Probes Based On Rhodamine Dye And Their Application In Biomedical Imaging

In the field of biomedicine,compared with traditional analysis techniques,fluorescent probe analysis technology has the advantages of real-time detection,less time-consuming and low cost.More and more people love it.There are many common dyes used as fluorescent probes,and rhodamine is one of them.Rhodamine and its derivatives have the characteristics of strong fluorescence signal,long wavelength and less damage to tissues.Therefore,the main research content of this article is to use rhodamine dyes to modify them at specific sites to synthesize some fluorescent probes with excellent properties that can detect biological molecules and ions.There is a complex relationship between small active molecules in cells.At the level of organelles,active small molecules play an important role in maintaining the function of organelles.In order to study the relationship of molecules in subcellular components,it is necessary and important to develop a detection tool that can track two or more related biomolecules in the organelle and emit multiple fluorescent signals.At present,there are very few probes that meet this requirement,which is still a difficult challenge to break.Here,we propose a novel single fluorescent probe?Lyso-HA-HS?,which can detect oxidizing substances?HOCl?and reducing substances?H₂S?in organelles?lysosomes?.The reaction of the Lyso-HA-HS probe with HOCl and H₂S produces different signal responses to oxidative and reducing substances in the lysosome,which are displayed in the red and blue channels of the microscopic imager,respectively.At the same time,for the hypochlorous acid and hydrogen sulfide produced by intracellular metabolism,the synthesized Lyso-HA-HS fluorescent probe can be used for dual-channel imaging in the lysosome using a confocal microscope.In addition,the excellent characteristics of the probe Lyso-HA-HS such as high selectivity,good membrane permeability and lysosomal enrichment ability can be used to reveal the relationship between HOCl and H₂S in lysosomes.Although palladium ions are low in organisms,they are enriched in organisms and cause a great deal of damage to them.Therefore,it is very necessary and important to develop a molecular tool capable of proportional imaging of Pd²⁺in living cells.Here,we developed a new fluorescence probe based on FRET mechanism?CR-Pd?,and it was proved experimentally that the probe only reacts to the divalent palladium ions,while there is little reaction to other valence palladium ions.The CR-Pd probe introduces two fluorescent groups,which act as energy donors and energy acceptors,respectively.The probe can specifically measure the Pd²⁺content in living cells.The cell experiment results show that when only the probe molecule is added to the cell,only the blue light generated by coumarin under the confocal microscope.When the palladium ion is combined with CR-Pd,the rhodamine carbonyl part is complexed with the palladium ion.The charge rearranges,the spironolactone ring is opened,the blue fluorescence decreases and the red fluorescence appears.At the same time,CR-Pd can be used as a probe to visualize the ratio of Pd²⁺in mitochondria.The small molecules and ions in the living body are in dynamic balance.If the content of any molecule is too high or too low,it will cause the disorder of the body's dynamic balance.Therefore,if the content of HCl O in the cell is too high or too low,it will damage the tissues of the organism and induce various diseases^[1].Based on the above,we have developed a fluorescent probe CR-HA that can detect HCl O,the synthesis steps of this probe are simple.By calculating the relationship between the ratio of the fluorescence intensity of coumarin and rhodamine and the concentration,a fitting curve is obtained to determine the content of HClO.This method can avoid many errors,such as the concentration and excitation of the probe.Factors such as strength interfere with the test results.At the same time,the probe has a two-photon property,which avoids tissue damage to the sample to be measured due to the short excitation wavelength,and largely avoids damage to biological cells during imaging.Spectral experiment results show that the probe has good selectivity,a wide range of acid and alkali applications,and a rapid reaction with HClO.Fluorescence imaging results show that the CR-HA probe is used to detect HClO in living cells,and the CR-HA probe can also detect HCl O in living tissue and zebrafish.