الفهرس | Only 14 pages are availabe for public view |
Abstract In this thesis, we studied binding of [Ru(NO)(Et2NpyS4)]Br complex (RuNOTSP) and its ligand (TSP) as drugs with biomolecules BSA and DNA using UV-Vis, fluorescence spectroscopy, Stopped-flow and molecular docking. The thesis consists of three chapters as follows: Chapter 1 provides a general information about ruthenium metal and their complexes, ligand substitution, the kinetics of ruthenium compounds, and different mechanisms of action. This gives them advantages over platinum- based complexes. Then, we discuss the application of ruthenium in diverse fields along with its physical and chemical properties. We focus on the biomedical application which divided into treatment and diagnostic aspects. Ruthenium and their complexes have anticancer, antimicrobial and immunosuppressant activity. Also, they are used in determination of cyclosporine, ferritin and folate level in human body. This study has focused attention on the interaction between ruthenium nitrosyl complexes and BSA and DNA as biological targets. Also, this chapter outlined importance of serum albumin and deoxyribonucleic acid. Chapter 2 includes the experimental details of the current study, which includes: i) The material used in the current work; ii) Synthesis of ligand and ruthenium nitrosyl complex; iii) Instruments and equipment. Also, details of the biological investigations as Serum albumins and deoxyribonucleic acid binding measurements, antibacterial activity and cytotoxicity. In addition, spectroscopic technique, kinetic experiments details and computational studies are described. Chapter 3 presents results and discussion. Our findings can be summarized as follows: Nitrosyl ruthenium complex RuNOTSP were synthesized in our lab as reported previously via template alkylation of Bu4N[Ru(NO)(S2C6H4)2] with Et2Npy(CH2Br)2 in boiling THF. RuNOTSP was characterized using UV-Vis, IR, Mass spectroscopies conductivity measurements and compared with published data. The interaction of the nitrosyl ruthenium complex RuNOTSP and its ligand TSP with BSA were studied using stopped-flow, fluorescence quenching, UV-Vis absorption studies, and molecular docking. Kinetic studies of interaction of ligand and its complex with BSA using stopped-flow can be summarized as follow: a first fast second-order binding and reversible step of BSA binding including complex dissociation and formation was suggested for all compounds. For the second reaction steps, a slow first-order isomerization reaction, a reversible reaction step was observed for TSP whereas, an irreversible reaction with RuNOTSP indicates that ligand-BSA complex is more stable kinetically than complex-BSA complex. The ?G values for the overall reaction are -10.5 kJ mol-1 for RuNOTSP and -7.4 kJ mol-1 for RuNOTSP and TSP indicated the reaction is spontaneous for both compounds. Fluorescence quenching of both compounds observed the presence of the static quenching mechanism through complex formation. The quenching constant (Ksv) of RuNOTSP calculated as 1.5 × 105 M-1 at 298 K is higher than the corresponding Ksv of TSP, which is 9.4 × 104 M-1 that indicates high affinity of the ligand towards BSA. TSP and its complex were checked for their antibacterial activity against E. coli, P. aeruginosa, S. aureus and E. faecalis bacteria where important activities were obtained with a good relationship to the coordination affinity and the binding constants. The cytotoxic activity of both compounds was tested against Hepatocellular carcinoma (HepG2) and normal liver cell lines (BNL) at different concentrations by WST-1 assay. The results showed that the survival of tumor cells increased with increasing concentration and the cytotoxicity trend found Ru-NO complex is higher than ligand. The molecular docking study illustrated that van der Waal, Pi-Alkyl interaction, and the hydrogen bonding play an important role in BSA binding. Also, the interaction of DNA with RuNOTSP and its ligand TSP are studied by fluorescence quenching, UV-Vis absorption, stopped-flow, and molecular docking. Fluorescence enhancement spectra of both compounds is controlled by a static enhancement mechanism. The dynamic constant (KD) of RuNOTSP calculated as 2.4 × 104 M-1 at 298 K is slight lower than the corresponding KD of TSP, which is 2.7 × 104 M-1. The absorption spectra of RuNOTSP and TSP decrease with increasing concentration of DNA. The binding constant (kb) of RuNOTSP calculated as 4.05 × 104 L.mol-1 when TSP is 1.77 × 104 L.mol-1. Kinetic studies of interaction of RuNOTSP and TSP with DNA using stopped-flow via two different mechanisms, a fast reversible followed by slow reversible steps were observed for RuNOTSP and TSP. The optimized geometric structures of ligand and its metal complexes are in good agreement with the experimental results. Molecular docking shows that RuNOTSP and TSP interact in a parallel manner with the major groove of DNA backbone through non-covalent interaction such as hydrogen bonding, van der Waals and hydrophobic interactions. The obtained data indicate that RuNOTSP has a higher activity than TSP. |