Electrochemical Studies Of Interaction Of Porphrins And Other Drugs With DNA At Interface

The interaction of drugs with biological molecules has been studied, which can help us understand function mechanism of drugs on the molecule level, and provide us with valuable information to design new drugs. Acting as the carrier of genetic information, DNA plays an especially important role in the course of genetic, such as memory, compilation, transfer and expression in cells, and expedites and controls the course of metabolism. DNA is the main target molecule that exists in anticancer and antiviral therapies. All cells and virus carry DNA. Normal heredity function of DNA depends on the physical and chemical interaction of other molecules with DNA.Interaction of DNA with small molecules, especially drug molecules affected physiological and physical chemical character of DNA and changed the transfer and copy of DNA. Hence it is meaning that interactions of small molecules with DNA are investigated to understand bind of DNA with protein, anticancer mechanism and to design and synthesize new drug. In medicament, it possible to foresee major steps forward in our understanding of the molecular basis of disease, both from attack by external pathogens and internally from variations within the human genome resulting in a plethora of new molecular therapeutic targets for drug design and discovery.A variety of techniques from molecular biology have been used in research laboratories to study the effects of DNA binding drugs, such as the gel mobility shift assay, H NMR, DNA foot-printing assay, and fluorescence-based assays, UV-Vis spectrum and viscometry. However, most of these methods are indirect and discontinuous, and require various labeling strategies. Moreover, the turnover time and cost of the conventional assays place a limit on the applicability of large-scale screening of drug candidates. With the development of modern techniques of chemical synthesis, compound separation, chemical molecular structure analysis, and the theory and investigation techniques of molecular recognition, much attention are paid to the interaction of DNA with other molecules recently. DNA modified electrode has some advantages over the conventional method, such as less sample consumption, higher sensitivity and signal/noise ratio and so has made great progress in the field concerning DNA studies, especially construction of biosensor and gene chips. Molecular self-assembled monolayers(SAMs) modified have been widely investigated during the last two decades. However, SAMs modified electrodes have seldom been used as the substrate for adding antibiotics on it and form a novel multi-layer film.Based on the above view, in this paper the gold electrode modified insolvable porphyrin and its metal complexes self-assembled monolayer (SAM) was prepared, and the mechanisms of their interaction with DNA was studied at solid-liquid interface on gold electrode by cyclic voltammetry, scanning electrochemical microscope (SECM), and alternating current (AC) impedance. The electrochemical studies of interaction between DNA and anticancer drugs, the electrochemical property and their application of the DNA modified monolayers electrodes by self-assembling also have been probed, and some new results has been obtained.There are five parts in this paper. Main contents are as follows:Chapter I General Introduction and ConclusionIn this part, the development in the electrochemical studies of DNA, including electrochemical behavior of DNA, electrochemical analysis of DNA, interaction of DNA with other matter and DNA modified electrode. It is important to study DNA interaction with drug in theory and in practice. self-assembled monolayer (SAM) modified electrode and the development about chemically modified electrodes including the theory, study methods and application in the area of analytical chemistry is presented. Ideas and purpose of the study is established.Chapter II The Preparation and Electrochemical Studies ofThiol-porphyrins (H₂MPTPP and H₂MBTPP) Self-assembled MonolayerModified Gold Electrode and Interactions with DNASince the first observation of the preferential accumulation of hematoporphyrin in neoplastic tissues by Policard in 1924, porphyrins and their derivatives have roused people's much greater concern. Water-insoluble and water-soluble porphyrinyl-nucleosides containing adenosine and thymidine, have strong tumoricidal activity against human malignant melanoma. The DNA-binding interactions of them have long been of interest because of the potential for therapeutic applications and the novel binding interactions observed with DNA. Porphyrin-DNA interactions have been extensively studied using a wide variety of techniques. In many studies, porphyrins which interact with DNA are almost water-soluble porphyrin, while the interaction of water-insoluble porphyrins with DNA have rarely seen in report. We provide the method of self-assemble monolayers (SAMs) of water -insoluble porphyrins on gold electrode to study the interaction of prophyrins and DNA at the electrode/solution interface, using a new measurement—the scanning electrochemical microscope (SECM), which is an electrochemical tool that can be used to study immobilized biomolecules and chemical or biological reactions at various interfaces with high spatial resolution, and can obtain rate constants of homogeneous and heterogeneous redox reactions precisely. We provide insight into the interaction of DNA with porphyrins and metalloporphyrin SAMs on gold electrode by means of CV, SECM, and A.C. impedance methods using a redox couple as a probe. After totally and partly modified Au electrode by H2MPTPP interacted with DNA, the results are similar: the decrease of current and loss of reversibility of the redox reaction for K₃[Fe(CN)₆], apparent heterogeneous reaction rate constant (k_eff from SECM and k_f from A.C. impedance) was decreased, and the hindrance (B) of electrode increased, which show DNA was adsorbed on the porphyrin and increased the inhibition of ET. These results prove that DNA can interact with H₂MPTPR From above results, we suggest that the mode of the interaction between H₂MPTPP and DNA is electrostatic interaction.Chapter III The Preparation and Electrochemical Studies ofMetal-porphyrins (CoMPTPP and NiMPTPP) Self-assembled MonolayerModified Gold Electrode and Interactions with DNASome metalloporphyrins usually play important roles in nature as mentioned previously. Investigation of the interaction of metalloporphyrins and DNA may lead to medical applications in the inhibition of the AIDS virus, and in the photodynamic treatment of tumors. So it is very significant area of research.In this work, we have carried out the cyclic voltammetry (CV), scanning electrochemical microscope (SECM), and A.C. impedance methods to study the interaction of metalloporphyrin (Ni, Co-MPTPP) with DNA and compared with porphyrin (H₂MPTPP). The difference from Co-MPTPP is that the current decreased after interacted with DNA, and the oxidation peak potential shifted (E_pa) negatively by 56mV and the reduction peak potential (E_pc) shifted positively by 70mV, but the redox peak currents of Ni-MPTPP interacted with DNA increased and redox peak potentials all shifted negatively by 80mV. The k_eff from SECM and k_f from A.C. impedance and the hindrance (B) of electrode were also detected, which show DNA was adsorbed on the porphyrin and increased the inhibition of ET. It is suggested to be electrostatic interaction of H₂MPTPP, Co-MPTPP and Ni -MPTPP with DNA, and the order of attractive force between porphyrins and DNA is Ni-MPTPP > Co-MPTPP > H₂MPTPP.Chapter IV The Preparation and Electrochemical Studies of DNA Self-assembled Monolayer Modified Gold ElectrodeWe have studied the micropatterning and characterization of the organic monolayers using cyclic voltammetry (CV), scanning electrochemical microscope (SECM), atom force microscope (AFM), and A.C. impedance and determined the electrochemical parameters, i.e., apparent reaction rate constant (k_f), and the coverage of electrode surface (θ). Using the high sensitivity of the electron transfer of ferricyanide to the modification of the gold surface with DNA, we selected this reaction as a probe to study the different modification stages at this modified electrode. SECM images obtained from bare and partially and totally modified electrodes showed very good resolution with different topography or null according to the extent of modification. Based on a comparison with the results from experiments, it can be obtained a reasonable agreement, which means a conjunction of these techniques.The DNA monolayer exhibited good stability and long lifetime. It can stand solution washing, hence is suitable for electrochemical studies. The voltammetric behavior of electro-active compounds, kanamycin and erythromycin at the DNA-modified gold electrode was explored. According to the electrochemical parameters, it can be concluded that both kanamycin and erythromycin interact with DNA through intercalation.Chapter V Electrochemical Behavior of Adriamycin and its Cu²⁺ complex and Interaction with DNAThe antitumor antibiotics daunomycin (DM) and adriamycin (ADM) are in wide clinical use for the treatment of various neoplastic diseases. ADM is an anthracycline antibiotic widely used as an antitumor agent. It binds to DNA by intercalation between base pairs and inhibits RNA transcription. Its activity against a broad spectrum of solid tumors has determined the importance of the studies on its interaction with DNA. Because the metal ions are not only relative to toxicity for the heart, but can enhance the damage for the anthracycline to cleave DNA and improve the active anticancer as well, the studies of interaction of metal ions and anthracycline with DNA are very significant, which can help us understand the existence state, interaction mode and toxicity mechanism and design the chemistry synthesize for nuclein acid enzyme.By electrochemical method to study the difference of the electrochemical behavior of ADM with or without DNA, it can be found the influence of DNA on the electron transfer process of ADM. In this part, we report here the studies on the binding of ADM and its Cu²⁺ complex to DNA by surface electrochemical methods, DNA modified electrode and UV-Vis spectrum. Using cyclic voltammetry (CV), double potential step chronocoulometry (DPSCC), scanning electrochemical microscope (SECM), and A.C. impedance and determined the electrochemical parameters. The results shown that ADM and ADM-Cu both can bind to DNA at DNA modified glass carbon electrode, and their binds to DNA by intercalation mode. However, the force between ADM-Cu and DNA is stronger than ADM. An intrinsic binding constant, K of (2.67±0.4)×10⁶ M^-1, and binding numbers (n) of 1 of ADM with DNA were obtained. For the complex and DNA-GCE system we obtain binding numbers (n of 2) of complex per DNA (bp) and a saturation coverage value (T_s) for ADM-Cu at dsDNA-GCE and other electrochemical parameters. By SECM and A.C. impedance, the k_eff from SECM and k_f from A.C. impedance were obtained. The results provide new insight into rational drug design and would lead us to further understanding of the interaction mechanism between antitumour drug and DNA.