The Activities Of The Copper Complexes On Escherichia Coli And Preparation Of Several Chemically Modified Electrodes

The researches of this paper can be divided into two parts:(1) Microcalorimetry study of the antibacterial activities of some Cu-phen-amino acid (phen=l,10-phenanthroline) complexes on Escherichia coli (E. coli) and the discussion of the antibacterial mechanisms;(2) The preparation of several chemically modified electrodes for ascorbic acid detection, detection of specific oligonucleotide sequence of Hepatitis C virus subtype lb and Catalase immobilization. The research for each section was showed as follows,(i)(Section2) Copper complexes:[Cu(phen)(L-Ser)(H2O)Cl](1),[Cu(phen)(Gly)(H2O)]Cl-3H2O (2),[Cu(phen)(L-Ala)(H2O)]ClH2O (3),[Cu(phen)(L-Phe)(H2O)]Cl-2.5H2O (4), Cu(phen)2Cl2-6H2O (5) were synthesized and characterized. The structure of1was characterized by X-ray crystallography and showed in a triclinic system with space group P1, a=6.8953(15) A, A,b=10.737(2) A, c=11.894(3) A, a=110.395(3)°,β=94.183(4)°and γ=100.540(3)°. The antibacterial activities on E. coli of these five copper complexes and CuCl2(6) were investigated by microcalorimetry. By analyzing the metabolic thermogenic data and curves, crucial parameters such as rate constant of bacterial growth (k), half inhibitory concentration (IC50), and generation time (tG) were obtained. All these copper complexes could stimulate the growth of the E. coli at their lower concentration. At their higher concentration they all showed antibacterial action. The inhibition on E. coli was5>1-2-3-4>6.(ii)(Section3) The antibacterial mechanisms of CuCl2(1), Cu(phen)2Cl2'6H2O (2),[Cu(phen)(Gly)(H2O)]Cl·3H2O (3),[Cu(phen)(L-Ser)(H2O)Cl](4)(phen=1,10-phenanthroline) on E. coli were investigated. The abnormal elevation of intracellular copper in E. coli was measured by inductively coupled plasma atomic emission spectroscopy (ICP-AES). DNA binding models and the binding constants of these complexes to calf thymus DNA (CT-DNA) were detected by UV-vis spectra. Meanwhile, the cleavage activities on pBR322DNA were studied by gel electrophoresis (GE). And the redox potentials of Cu(II)/Cu(I) couple were measured by cyclic voltammetry (CV) experiments, which were considered to be an influence factor of the DNA cleavage abilities. The results of these experiments can well illustrate the reason why the antibacterial efficiency on E. coli is2>3>4>1.(iii)(Section4) A new dinuclear copper salicylaldehyde-glycine Schiff base complex [Cu2(Sal-Gly)2(H2O)2] was synthesized and structurally characterized.[Cu2(Sal-Gly)2(H2O)?] was crystallized in the monoclinic system, P21/c space group. The molecule is a dinuclear complex, formed by two [Cu(Sal-Gly)(H2O)] units. The electropolymerization properties of the copper complex on glass carbon electrode were studied at different potential ranges. The electropolymerization happened when the high scan potential reached1.4V. The modified electrode exhibited good electrocatalytic oxidation properties to ascorbic acid and showed a sensitivity of22.9nAμM-1(R2=0.9998) and detection limit of0.39μM (S/N=3) in the amperometric determination of ascorbic acid. The designed determination method can be used to analyze the Vitamin C tablets.(iv)(Section5) A label-free electrochemical DNA sensor was designed for detection of specific oligonucleotide sequence of Hepatitis C virus subtype lb, in which the electrochemically active compound plays a dual role. Firstly, L-cysteine (L-Cys) was assembled on gold electrode (AuE) surface. Then [Co(phen)2Cl2]Cl reacted with the immobilized L-Cys to produce [Co(phen)2(L-Cys)].[Co(phen)2(L-Cys)] were immobilized on the AuE surface acting as anchors for probe DNA immobilization and also as electrochemical indicators. The hybridization events were monitored by Square-wave voltammetry (SWV). The designed DNA sensor showed good selectivity, which can be used for one-base mismatch discrimination. And the peak intensity was linear with the logarithmic value of the complementary target DNA concentration from1pM to1μM with a correlation coefficient of0.99.(v)(Section6) A new method for protein immobilization based on metal-ion chelation was reported. First, conductivity film poly(aniline-co-2-aminobenzoic acid)(PAOAA), which is able to conductive in neutral solution, was prepared by electrochemical deposition. The film was characterized by scanning electron microscopy (SEM) and energy-dispersive X-ray (EDS). Then Cu2+were adsorbed on the film as the chelating ion to immobilize Catalase (Cat). The molar amount of Cu in the polymer was0.49%measured by EDS. Differential Pulse Voltammetry (DPV) was used to research the immobilization of Cat. The results showed that as PAOAA film forming and Cat immobilizing on the electrode surface, the peak current in10mmol/L K3Fe(CN)6/K4Fe(CN)6(Fe(CN)63-/4-) decreased from121μA (bare GE) to92.6and71.8μA respectively. This indicated that PAOAA film and Cat on the electrode could impede the charge transfer. This conclusion is same as the result of electrochemical impedance spectroscopy (EIS) studying in the process of Cat immobilization. The catalytic activity of immobilized Cat for hydrogen peroxide was studied by Linear Sweep Voltammetry (LSV) method. Finally, Chronoamperometry (CA) as the detection method, the Cat-modified electrode was able to quantitatively detect hydrogen peroxide. Moreover the Cat-modified electrode could be reuseable.