Synthesis Of Functional Fullerene Derivatives And Their Application In Electrochemical DNA Sensors

The pristine fullerene (C60) has a closed cage structure, which greatly limits its application in many areas of life science, drug development and materials science. In order to overcome this obstacle, fullerene has been chemically functionalized by covalent attachment of different groups on its globular framework to alter the physical and chemical properties of fullerene. In this dissertation, novel fullerene derivatives were synthesized by C60 1,3-dipolar cycloaddition and hydroxylation reaction and applied to the electrochemical DNA sensors. The main contents are as follow:(1) A new C6o pyrrolidine derivative was synthesized by the 1,3-dipolar cycloaddition reaction of C6o with o-vanillin and glycine. The effects of the molar ratio of the reactants and the reaction temperature on the reaction were investigated respectively. The optimal reaction condition was that C6o, o-vanillin and glycine with the molar ratio of 1:2:5 in toluene were refluxed at 100℃ for 24 h under a nitrogen atmosphere. Then crude product was separated and purified by silica gel column chromatography to get the C6o pyrrolidine derivative (2-(2-Hydroxy-3-Methoxyphenyl) [60] Fulleropyrrolidine). The structure was characterized by high performance liquid-mass spectrometry (HPLC-MS),1H nuclear magnetic resonance (’H NMR), infrared (FT-IR) and ultraviolet-visible absorption spectroscopy (UV-Vis). The morphology and spectral properties of product were detected by atomic force microscopy (AFM) and fluorescence spectra (FS), respectively.(2) 2-(2-hydroxy-3-formyl-phenyl)-5-(hydroxymethyl) C60 pyrrolidine derivatives (FPD) were successfully synthesized, separated and purified by the reaction of C60 with o-vanillin and L-serine. The structure was characterized by elemental analysis (EA), HPLC-MS,’H NMR, FT-IR and UV-Vis to determine its molecular formula was C70H13NO3, and the fluorescence and electrochemical properties of the product were investigated.(3) FPD was immobilized on the surface of glassy carbon electrode (GCE) by the direct coating method, and then an Al (III) film was assembled on its surface. The probe DNA was fixed on the electrode surface through the electrostatic interaction between phosphate group of DNA and aluminum (III), and an electrochemical DNA sensor based on A13+/FPD/GCE modified electrode was constructed successfully. The analytical performance of the biosensor was detected through using methylene blue (MB) as the hybridization indicator. The peak currents of MB showed good linear relationship with the logarithm value of the target concentrations (lgCs2) in the range from 1.0×10-15 to 1.0×10-9 mol/L, with a linear equation of Ip/10-6 A= 0.106 lg(CS2. mol/L)+0.735, and a limit of detection of 5.4×10-16 mol/L. The selectivity experiments further showed that the complementary sequences, non-complementary sequences and the base mismatch sequences could be well distinguished by this biosensor.(4) Water-soluble fullerenols was prepared in accordance with the best experimental program of the research group. Fixed ssDNA with sulfydryl on the gold electrode surface and constructed a novel DNA sensor by use of the specific affinity interaction between fullerenols and single-stranded DNA. The oligonucleotides related to CaMV35S promoter fragment were determined with the developed biosensor via electrochemical impedance spectroscopy (EIS) using [Fe(CN)6]3-/4- as indicating probe. A linear range of 1.0×10-16 to 1.0×10-9 mol/L and a limit of detection of 2.0×10-17 mol/L were obtained. The selectivity experiments showed that the biosensor could identify the complementary sequence, non-complementary sequence and the base mismatch sequences effectively.