Design, Synthesis And Properties Of Metal Complex Luminescent Probes And Nucleic Acid Sensors

It is of important significance to develop rapid, sensitive and cost-effective methods for the detection of nucleic acid in the fundamental biological/biomedical research. Biosensors relying on fluorescence in DNA detection, have attracted great attention because of their ultrasensitivity, relatively rapid and easy operation approaches. Most classic DNA fluorescent dyes mainly bind DNA with secondary structures by intercalation or groove binding, and thus many of them do not show significant fluorescence response for ssDNA without secondary structures. To sense ssDNA effectively and conveniently, some new fluorescent dyes based on electrostatic interaction were developed. However, the practical applications of these ionic probes are limited, because electrostatic interaction is easily affected by other electrolytes. Alternately, metal-ligand coordination interaction has been widely used in anions recognition and sensing, and work more effectively than electrostatic interaction or hydrogen bond interaction.Based on these research, we incorporated coordination interaction to design sensors for DNA detection. A series of new metal complexes were synthesized and tested. This thesis is divided into nine chapters as follows:1?Introduces the basic principles used in fluorescence sensing, various types of fluorescent reporters and recognition units. The focuses are anions sensors, nucleic acid probes and aptamers. The design strategies and main contents of the thesis are outlined.2?A fluorescent probe based on pyrene and Zn(?)-dipicolylamine unit, PyZn, was designed and synthesized for anions and DNA detection. The emission of PyZn decreased significantly with increased water fractions in ethanol/water solutions. Under the optimized condition, PyZn showed enhanced excimer emission and reduced mono emission upon the addition of DNA in HEPES buffer solutions, which revealed that Zn(?)-dipicolylamine unit binds DNA by coordination interaction and induce the aggregation of pyrene.3?A new fluorescent indicator-displacement assay for anions was developed by using two zinc complexes based on9,10-distyrylanthracene, DSAZnl and DSAZn2. The fluorescence of the two complexes can be quenched significantly upon the binding of a common indicator--phenol red (PR) in HEPES buffer solutions. And the subsequent addition of citrate, pyrophosphate (PPi) or DNA to this solution can recover the fluorescence to varying degrees due to PR displacement by analytes coordinating to the complexes. This approach showed good selectivity over other eleven anions. By using10?M DSAZn2and PR as complexes and indicator respectively, a linear response for citrate detection was observed in the range0-10?M with a detection limit of0.1?M. Different binding modes of the two complexes to various analytes were dicussed in terms of fluorescence and absorption titration spectra.4?By taking advantage of the coordination interaction and the emission of restricted tetraphenylethene, two zinc complexes, TPE4Zn and TPE1Zn were designed and synthesized by modifying tetraphenylethene with Zn(II)-dipicolylamine unit. To demonstrate the differences between coordination and electrostatic interactions, tetraalkylammonium moieties functionalized tetraphenylethene, TTAPE, was synthesized for comparison. TPE4Zn showed much higher sensitivity in detecting DNA than TTAPE in aqueous solutions. And the fluorescence enhancement of TPE4Zn was not depend on the length of DNA, which proved that the coordination interaction is more effective than electrostatic interaction. Compare to TPE4Zn, TPE1Zn showed much weaker self fluorescence in buffer solution, which maked TPE1Zn an ideal probe for DNA detection in aqueous medium. TPE1Zn exhibited much higher sensitivity in detecting both ssDNA and dsDNA than the classic intercalation dye of ethidium bromide.5?By using cis and trans tetraphenylethene derivatives as precursors, pure cis and trans binuclear Zn complexes, TPE2Zn, were synthesized and tested respectively. They exhibited weaker self fluorescence and higher sensitivity in detecting DNA than TPE1Zn. Moreover, cis complex showed significantly higher sensitivity and better selectivity than its trans isomer. In particularly for titration experiments with pyrophosphate, cis complex showed neglectable fluorescence change while trans complex displayed significant fluorescence enhancement. To our knowledge, this work represents the first report to address the difference of cis and trans configuration isomer in affinity of binding analytes.6?By using cyclen as starting material, a tetraphenylethene functionalized Eu(III) macrocyclic complex, TPEEu, was synthesized for protein and DNA detection. TPEEu exhibited dual emission of tetraphenylethene fluorescence and Eu(III) phosphorescence with a low quantum yield. This dual emission can be significantly emhanced upon the addition of BSA and DNA, which revealed that the restricted tetraphenylethene can act as an antenna and transfer its excited state energies to the lanthanide excited state. This probe based on Eu(III) phosphorescence emission can be easily distinguishable from fluorescence background emission or autofluorescence, and greatly enhance the signal-to-noise ratio by using time-resolved techniques.7?A novel label-free method for DNA-based sensors was developed by using a minor groove binding dye-DAPI. By using different DNA sequences, we developed turn off and turn on sensors for Hg2+detection with high sensitivity and selectivity. The detection limits were5nM and1.5nM for turn off and turn on sensors respectively, which is lower than the U.S. Environmental Protection Agency (EPA) standard for the maximum allowable level of inorganic Hg(II) in drinking water. Compare to those sensors needing covalent labelling, modification or immobilization, the label-free strategy has advantages in operation convenience and cost. We demonstrate that the nucleic acids can not only non-covalently bind fluorophores to sense targets, but also display opposite fluorescence responses for the same targets by designing the specific sequences.8?The minor groove binding nucleic acids dyes, DAPI and Hoechst33258, were used in label-free aptamer-based sensor for L-argininamide. By using DAPI and Hoechst33258, the detection limit were3.8?M and2.5?M respectively. Both sensors showed good sensitivity toward L-ArgNH2over L-arginine and glycine. This label-free approach can be further extended to other aptamer-ligand systems for detecting and quantifying a broad range of analytes.