Synthesis Of Three Kinds Of Fullerene Pyrrolidine Functional Derivatives And Their Application In Electrochemical Biosensor

Fullerene (C60) has uniquely physical and chemical properties, which make it revealing broad application prospects in the frontier research fields of physics, macromolecule materials and life science. However, because fullerene possesses closed cage structure without contain any other active groups, its application scope is greatly limited. Cycloaddition reaction is one of the most important methods of the chemical modification of fullerene. It has been widely used to prepare the functionalized fullerene derivatives. This dissertation focuses on the researches on the synthesis, separation and characterization of functionalized fulleropyrrolidines, which were applied to the fabrication of electrochemical biosensor. The detailed research content is as follows:(1) With C60, terephthalaldehyde and glycine as resourse,2-(4-Formylphenyl)[60]Fulleropyrrolidine (FPD Ⅰ) were synthesized through the1,3-dipole cycloaddition. The products were characterized by1HNMR, MSn (n=1,2), FTIR and UV-Vis, and its chemical formulas was C69H9NO. The optical and electrochemical properties of the derivative were also determined by fluorescence spectrum and differential pulse voltammetry.(2) The product FPD Ⅰ was modified on the surface of glassy carbon electrode (GCE) by the direct coating method. The Human Chorionic Gonadotropin antibody (Ab-HCG) was then covalently bonded onto the surface of electrode through an amide condensation reaction between aldehyde group and amine group, and thus a novel electrochemical immunosensor based on fulleropyrrolidines was fabricated. By the biosensing technology combining the specifity of immune reaction and the high sensitivity of electrochemical impedance spectroscopy (EIS), the quantitative detection ability of the proposal immunosensor towards Human Chorionic Gonadotropin (HCG) were performed, and a linear range of0.1～10μg/mL and a detection limit of0.028μg/mL were obtained.(3) A compound of2-(2-Hydroxy-3-Methoxyphenyl)[60] Fulleropyrrolidine (FPD Ⅱ) was synthesized by the reaction of the C60and azomethine ylide (prepared from glycine and o-vanillin). FPD Ⅱ was immobilized on the surface of GCE by the direct coating method. By utilizing the favourable coordination of zirconium (Ⅳ) towards oxygen-containing groups (phosphate group), the probe DNA related to CaMV35S promoter gene on the electrode surface. The target NDA promoter fragment were determined with the developed biosensor via using [Fe(CN)6]3-/4-as probe. A linear range of1.0×10-13～1.0×10-9mol/L and a detection limit of4×10-14mol/L were obtained. The experimental results demonstrated that the electrochemical biosensor have good selectivity to recognize the complementary sequences, base-mismatched sequences and non-complementary sequences.(4) A compound of2-(4-Formylphenyl)-5-(1-Mercaptomethy)[60] Fulleropyrrolidine (FPD Ⅲ) was synthesized by the1,3-dipolar cycloaddition reaction of C60with terephthalaldehyde and L-cysteine. Gold nanoparticles were prepared on the surface of the electrode by electrodeposition method. FPD III was then self-assembled on the surface of gold nanoparticles through the Au-S bond. By choosing fulleropyrrolidines as covalent platform of DNA, the amide condensation reaction between aldehyde group and amine group was applied to immobilize the probe DNA. A novel electrochemical DNA-biosensor was then developed. With the electroactive probe of [Ru(NH3)6]3+as hybrid indicator, the chronocoulometric method (CC) was used to the immobilization of probe DNA and its hybrid performance. According to the different amount of [Ru(NH3)6]3+caused by electrostatic adsorption with DNA sequences before and after hybridization, the charge difference (△Q) was resulted. The relationship between the of [Ru(NH3)6]3+and logarithmic of target DNA sequence concentrations (1gC) was linear in the concentration range from1.0×10-16to1.0×10-11mol/L, and the detection limit had been estimated as2.4×10-17mol/L. The experimental results showed that the electrochemical biosensor could effectively identify different target DNA sequences and has excellent stability and reproducibility.