Study Of DNA Electrochemical Biosensor Based On Polypyridyl Metal Complexes As Electrochemical Indicators

In this article, series of polypyridyl metals were synthesized, whose structures were characterized by IR and EA methods. Cyclic voltammetry (CV) and fluorescence spectroscopy techniques were used to study the interaction between these compounds and salmon sperm DNA to optimize conditions. These sensors were prepared by immobilizing single-stranded DNA probes on glass carbon electrodes (GCEs) and using different electroactive indicators to measure the hybridization events between the DNA probes and their complementary DNA fragments. The electrochemical DNA biosensor might have potential application in disease testing or novel anticancer drug designing. This masteral dissertation constitutes by three parts besides the preface.(1) The copper complex of 4,5-diazafluorene-9-one (dafone) and bromine ligands ([Cu(dafone)2]Br2) was prepared and its interaction with double-stranded salmon sperm DNA (dsDNA) in pH 8.0 Britton-Robinson (B-R) buffer solution was studied by electrochemical experiments at GCE surface. The experimental conditions for incubation of dsDNA and [Cu(dafone)2]2+ such as pH, reaction time, scan rate and dsDNA concentrations were optimized to pursue the maximum anodic peak current difference. It was revealed that Cu(dafone)2Br2 could bind with salmon sperm DNA strands mainly by intercalation mode. The binding number of [Cu(dafone)2]Br2 for each salmon sperm dsDNA chain and equilibrium constant of the binding reaction were calculated to be 3 and 2.80×1012 L3·mol-3, respectively. The Cu(dafone)2Br2 was further utilized as a new electrochemical DNA indicator for the detection of DNA fragment. The difference of its electrochemical responses occurred between hybridized dsDNA duplex and probe DNA was explored to assess the selectivity of the developed electrochemical DNA biosensor. The constructed electrochemical DNA biosensor achieved a detection limit of 3.18×10-9 mol·L-1 for complementary target DNA.(2) A new Mn(II) complex of MnL2Cl2 (L= Di[azino-di(5,6-azafluorene)-К2-NN']) was synthesized and utilized as an electrochemical indicator for the determination of hepatitis B virus (HBV) based on its interaction with MnL2Cl2. The electrochemical behavior of interaction of [MnL2]2+ with salmon sperm DNA was investigated on GCE, and its interaction mode was found to be primary intercalation binding[0]. The extent of hybridization was evaluated on the basis of the difference between signals of [MnL2]2+ with probe DNA before and after hybridization with complementary sequence. Control experiments performed with non-complementary and mismatch sequence demonstrated the good selectivity of the biosensor. With this approach, a sequence of the HBV could be quantified over the range from 1.76×10-8 mol·L?1 to 1.07×10-6 mol·L?1, with a detection limit of 6.80×10-9 mol·L?1.(3) The electrochemical behavior of [CuL2]2+ on binding to DNA at a GCE was investigated on GCE. CV coupled with fluorescence spectroscopy techniques were used to study the interaction between [CuL2]2+ and salmon sperm DNA. Results showed that this system has two types of binding mode in the molar ratio range studied. An electrochemical DNA biosensor was developed using [CuL2]2+ as an electrochemical indicator. Numerous factors affecting the probe immobilization, target hybridization, and indicator binding reactions were optimized to maximize the sensitivity and speed the assay time. Using this approach, complementary target sequences of HBV can be quantified over the range of 1.74×10-9 ~ 3.45×10-7 mol·L?1 with a detection limit of 8.32×10-10 mol·L?1.