Design,Synthesis And Application Of Rhodamine-based Fluorescent Probes In Metal Ion Detection

Excessive accumulation of metal ions in living organisms can cause serious health problems.We find ways to detect metal ions in living organisms efficiently.Traditional detection methods have many shortcomings,such as extended response time and poor specificity,and cannot be applied in biological imaging.Fluorescence imaging technology has the advantages of high sensitivity and fast analysis speed.It is a tool that can visually monitor metal ions in the environment.This technology is highly dependent on fluorescent probes’photophysical and photochemical properties.Rhodamine-based fluorescent probes have a unique spiro-ring structure and excellent luminescence properties.A series of derivatives prepared based on them are valid for constructing switch-type fluorescent probes.It is favored in the preparation of biological fluorescent molecular probes.In this paper,three rhodamine-based fluorescent probes for detecting metal ions were constructed based on fluorescence analysis technology.Its spectral properties were characterized by experiments such as ultraviolet and fluorescence tests.At the same time,its potential application value is proved by studying actual water samples and the fluorescence imaging of living cells,animals,and plants.（1）A fluorescent probe rhodamine pyridinediaminethiophene（RBKB）,which responds explicitly to Al³⁺,was synthesized using rhodamine B as the fluorescent precursor 2,6-aminopyridine as the linking arm,and 2-thiophene acetyl chloride as the ligand.The structure of probe RBKB was confirmed by ¹H NMR,¹³C NMR,and ESI-MS.Ultraviolet-visible and fluorescence emission spectroscopy verified that it could detect Al³⁺with high sensitivity and selectivity in Et OH/H₂O（1:1,v/v）system solution without interference from other cations and anions.Job’s plot experiment showed that the binding ratio of probe RBKB to Al³⁺was 1:1,and the binding constant was 3.69×10³ M^-1,the detection limit for Al³⁺was 0.177μM.The recovery rate of probe RBKB to Al³⁺in actual environmental water samples was 90.5%-96.8%.At the same time,it was successfully detected Al³⁺in breast cancer cells（MCF-7）,zebrafish,and soybean due to its good water solubility and low toxicity.（2）The probe rhodamine pyridinediamine p-cyanobenzene（RBGP）was synthesized by using rhodamine B as the fluorescent precursor,2,6-aminopyridine as the linking arm,and 4-cyanobenzoyl chloride as the ligand.The structure of probe RBGP was confirmed by ¹H NMR,¹³C NMR,and ESI-MS.UV-Vis and fluorescence spectra show that it can respond rapidly to Al³⁺in Et OH/H₂O（1:3,v/v）system solution without interference from other cations and anions.Job’s plot experiment showed that the binding ratio of probe RBGP to Al³⁺was 1:1,the binding constant was 1.01×10⁴ M^-1,and the detection limit was 0.146μM.The addition of F^-could realize the reversible fluorescence response of the probe to Al³⁺.Compared with the previous probe RBKB,the probe RBGP has a shorter response time,lower detection limit,and higher binding constant.Experiments show that the probe RBGP can detect Al³⁺in actual environmental water samples.At the same time,it can successfully detect Al³⁺in breast cancer cells（MCF-7）,zebrafish,and soybean root cells in fluorescence imaging due to its good water solubility and low toxicity.（3）A rhodamine-based near-infrared fluorescent probe rhodamine-coumarin phenylhydrazine（RBLY）was synthesized by coumarin and ketoacid.Compared with traditional probes,near-infrared（650-900 nm）fluorescent probes have a more comprehensive working wavelength range and minor light damage to biological samples.They can more effectively avoid the interference of biological tissue autofluorescence during biological imaging.The structure of the probe RBLY was confirmed by ¹H NMR,¹³C NMR,and ESI-MS.UV-Vis and fluorescence spectra indicated that it could respond rapidly to Hg²⁺in Et OH/H₂O（1:5,v/v）solution without interference from other cations and anions.The fluorescence emission spectrum shows that its absorption peak is 620 nm and its emission peak is 695 nm.The saturation response time of the probe RBLY and Hg²⁺was 120 s,the binding ratio was 1:1,the binding constant was 1.63×10⁵M^-1,and the detection limit was 0.34μM.The addition of S^2-could make the probe show a reversible effect.The probe RBLY has a high recovery rate of Hg²⁺in actual environmental water samples.At the same time,it has good biocompatibility and can successfully detect Hg²⁺in the fluorescence imaging of MCF-7 cells,zebrafish,and soybean roots.