Noninvasive Study Of Intracellular Phosphorous Metabolism And Free Mg～(2+) By NMR

Nuclear magnetic resonance (NMR), as a spectroscopic technique, has been unprecedentedly developed in its applications to biomedical systems. The aspects of its applications involve structure of biopolymers in solution, in vivo NMR spectroscopy and magnetic resonance imaging. Its applications also involve the determination of free Mg²⁺ and the pH in several kinds of blood platelet and cells. (31)^P and (19)^F NMR methods used in determining intracellular free Mg²⁺ have provided much information for biological analysis. NMR, a noninvasive method, can study free ions and metabolism in the cultured cells simultaneously. (31)^P NMR was proverbially applied in perfused organs, experimental animals and human. In this thesis, cos-7 cells, rabbit blood platelet, human blood platelet have been studied by using (31)^P NMR, and pH values in these samples have been estimated. 5F-APTRA, an indicator used in (19)^F NMR method, also has been synthesized. This thesis includes four chapters. Chapter one: The applications of NMR in studying free ions and phosphorous metabolism in biologic samples were reviewed. Chapter two: (31)^P NMR was used in the research of some cells and platelets samples for the determination of pH and the study of phosphorous metabolism. Cos-7 cells were gathered and centrifuged, then diluted with sodium chloride (0.9%) to 108cell/mL sample. This dense cells suspension was poured into sample tube and was observed by (31)^P NMR. Analyzing the spectrogram, intracellular pH was estimated according to the chemical shift of Pi. Fresh rabbit blood was obtained form New Zealand rabbit and was centrifuged to get fresh blood platelet. Human blood platelet was taken from hospital. Both kinds of platelets were determined by (31)^P NMR method and phosphorous metabolism was investigated. Cos-7 cells, rabbit blood platelet, human blood platelet have been studied by (31)^P NMR. The intracellular pH values of cos-7 cells and rabbit blood platelet are 5.58 and 5.39, respectively. Chapter three: (31)^P NMR method used in free Mg²⁺ determination has been established. ATP in cells combined with free Mg²⁺ and the chemical shift difference of α, βpeak will change with concentration of free Mg²⁺. We simulated intracellular ions intensity and acidity, and under the simulated condition KD^MgATP of MgATP was computed. So concentration of free Mg²⁺ was obtained indirectly.