Synthesis And Biological Application Of Fluorescein Dyes With Red Emission

Fluorescein is a clinically approved fluorescent dye with low toxicity and is commonly used as a fluorescent probe and fluorescent imaging reagent in the biomedical field.However,the green fluorescence of fluorescein itself,with short wavelength and high background interference,limits its biological applications.Therefore,this paper investigates its application studies in fluorescent probes and photodynamic therapy based on the red fluorescein-derived dyes DCF-MPYM and FL previously reported by our group.In this paper,the red fluorescein dye DCF-MPYM with a large Stokes shift was used as the parent,and the allyl ester group was introduced into its phenolic hydroxyl group as the recognition group of the divalent palladium ion to obtain the fluorescent probe DCF-MPYM-Pd.When the probe was not combined with the divalent palladium ion,the fluorescence was weak because the spiro-pyran was in the closed-loop state,and after the probe was combined with the divalent palladium ion,the open-loop fluorescence of spiro-pyran was turned on to achieve The purpose of detection.Importantly,the probe can be excited by green light and detected in the red light region,achieving low background interference,high selectivity,high sensitivity and specificity for the detection of divalent palladium ions,with a detection limit of 37.08 n M well below the drug threshold set by the European Council for the Quality Control of Medicines.Since DCF-MPYM can enter cells and has the ability to lysosomal localization,we performed cellular imaging of the probe and found that the probe also responds well to divalent palladium ions in cells and aggregates at lysosomes.The photosensitizer FL-Bio-V1 was obtained by linking another red fluorescein dye FL to biotin via hexamethylene diamine as a parent.By comparing the absorption and emission spectra of FL-Bio-V1 with FL,a good overlap was found,indicating that its photophysical properties did not change after linking to biotin.The ability of FL-Bio-V1 to produce both singlet oxygen and superoxide anion was verified by spectroscopic and cellular experiments,suggesting that FL-Bio-V1 can achieve a combined type I and type II PDT treatment.The photodynamic therapeutic effect of FL-Bio-V1 was demonstrated by cytotoxicity assay(MTT)and live-dead cell staining assay.The mechanism of FL-Bio-V1 photodynamic therapy was demonstrated to induce apoptosis by disrupting lysosomal integrity through acridine orange(AO)verification lysosome integrity assay.