Design, Synthesis And Sensing Of Optical Molecular Probes

Molecular probes have been gained great concern in the application ofenvironment and life science because of its characteristics, such as high sensitivity,numerous detection objects and rich response signal et al. Design and synthesis ofnovel molecular probes with excellent performances have become one of researchhots in the across-field between organic chemistry and analytical chemistry. For thedesign of molecular probes, the key factors that affect the performance of probes aremolecular recognition and signal transduction. In this thesis, we focus on thedevelopment of novel optical molecular probe. We chose spirobenzopyran andcyanine which have good optical performance as the skeletons, designed andsynthesized a series of functional molecular probes to detect metals ions, anions andbiological molecules with physiological species quantificationally, based oncolorimetric and fluorescent analysis methods. The thesis has done the followingworks:（I） Spiropyan probes for colorimetric detection of thiol-containing amino acidsand fluorinion based on chemical reaction.Two novel spiropyran derivatives were designed and synthesized, which wereapplied to detect neutral molecule and anion by its structure conversion.（1） We designed and synthesized a new spiropyran6,8-dinitro-1′,3′,3′-trimethylspiro [2H-1-benzopyran-2,2′-indoline]（DNSP） to and its application forrapid and sensitive colorimetric detection of thiol-containing amino acids in aqueoussolution, and proposed a concept-new mechanism for thermal switching of aspiropyran that is based on simultaneous nucleophilic-substitution reaction andelectrostatic interaction. The nucleophilic-substitution at spiro-carbon atom of aspiropyran is promoted due to electron-deficient interaction induced by6-and8-nitrogroups, which is responsible for the isomerization of the spiropyran by interactingwith thiol-containing amino acids. Further, the electrostatic interaction between thezwitterionic merocyanine and the amino acids would accelerate the structuralconversion. Compared with other reported spiropyran probes for amino acids, thiscooperative action improved the detection sensitivity with a detection limit of1.0×108M. The process can be considered that thiol-containing amino acids play therole as a mimetic enzyme. The activity of mimetic enzyme was discussed. （2） Based on the unique nucleophilic substitution reaction between fluorinionand silicon ether, we designed and synthesized a6-（tert-butyldimethylsilyloxy）-1’,3’,3’-trimethylspiro[chromene-2,2’-indoline]（SPS） which is colorless inCH3CN/Tris-HCl buffer solution, and no significant absorption is observed in thevisible region which suggests SPS is existed in the form of ring closure. In thepresence of F-, F-cleaves the Si-O bond incorporated in SPS which makes thespiropyran converts to its form of ring opening and the absorption at495nm isobserved accompanying with the colour chage from colorless to orange-red. So weconstracted a platform for recognizing and sensing F-, and developed a newmeachanism to regulate the optical character of spiropyran.（II） Indocyanine probes for fluorescent detection of Cys/and ascorbic acid basedon chemical reaction.Two novel near-infrared indocyanine derivatives were designed and synthesized,which were applied to detect neutral biological molecules having biological activity.Characteristic of near-infrared could effectively avoid some interference caused bybiological environment.（1） Aldehyde as the recognition group and thermodynamic reaction as guidingideology, a ratiometric fluorescent probe Cy5-A for Cys/Hcy was designed andsynthesized based on intramolecular charge transfer （ICT）. After reaction of probewith Cys/Hcy, the withdrawing-electron group of formyl became five-membered ringthiazoline or six-membered ring thiazinane, which weakened the pull electron abilityof formyl group, leading to the maximal emission peak red-shift about40nm. So theprobe could detect Cys/Hcy by ratio. Moreover, Cy5-A was successfully applied tothe quantitative detection of Cys in the fetal calf serum.（2） Based on that hydrazide could be hydrolyzed by Cu²⁺, a novel near-infraredindocyanine probe Cy5-HD was designed and synthesized. In aqueous solution, Cu²⁺can effectively hydrolyze N-N bond to get amino which could quench theindocyanine’s fluorescence because of the photo induced electron transfer （PET）.However, due to the oxidation-reduction reaction between ascorbic acid and Cu²⁺,Cu²⁺was reduced to Cu+, which inhibited the hydrolysis reaction lead to fluorescencerecovery. When the sufficient amount of ascorbic acid in system could reduce all theamount of Cu²⁺, the fluorescence of the system is strong. While, if the amount ofascorbic acid in system could not reduce all the amount of Cu²⁺, the system wouldcontain probe molecule and the hydrolysis product, resulting in the fluorescencedecreased. （III） Design of dual-functional probes.Two novel single-molecular multianalyte probes were designed and synthesized.Probe would produce different signal readout to two analytes existed alone orsimultaneously, which realized the simultaneous detection of multi-component incomplicated system.（1） Based on that the spiro C-O bond is easy to be protonated in acidic condition,a novel pH-switching single-molecular multianalyte probe Rhod-SP was designedand synthesized, which could colorimetric assay of Co²⁺and fluorescence response ofCu²⁺via regulation. Under acidic condition, as spiropyran was protonated to getphenolic hydroxyl, the binding constant between Rhod-SP with Cu²⁺is far greaterthan Co²⁺, triggering the ring opening of its spirolactam of rhodamine. However,Rhod-SP only selectively ligate Co²⁺under alkaline condition, resulting in the ringopening of spiropyran. So Rhod-SP could bind Cu²⁺reversibly by changing the pHcondition, which indicates that the sensor is an excellent candidate for exploring thegeneral pH environments of tumour and normal tissues. We demonstrated theapplicability for the fluorescent images of Cu²⁺in the breast cancer cell line （MCF-7）and normal breast cell line （MCF-10）.（2） Based on intramolecular photoinduced electron transfer and fluorescenceresonance energy transfer mechanisms, a dual-functional probe GCP was designedand synthesized, which was applied to detect intracellular β-D-glucosidase andphosphodiesterase I activity. GCP molecule possesses two cleavage sites, specificcorresponding to β-D-glucosidase and phosphodiesterase I, and three fluorescentreporters,7-β-D-glucopyranosyloxycoumarin,7-hydroxycoumarin and meso-tetraphenylporphyrin. Whether two enzymes exist only one or both, producesdifferent signal readouts with high sensitivity. Moreover, GCP is chemically stable incomplex biological fluids. Fluorescence outputs are not significantly affected bybiologically related metal ions, anions, amino acids and proteins. We demonstratedthe applicability for the simultaneous images of intracellular β-D-glucosidase andphosphodiesterase I activities by using the different optical imaging modes.This thesis proposed new design concepts and recognition mechanism fordeveloping novel optical molecular probes, and provided a guiding ideology forachieving multi-analytes detection in complex system.