Evaluation Of Affinity Interaction Between Cells And Small Molecules By Capillary Electrophoresis

Capillary electrophoresis(CE) has been widely used in evaluation of affinity interactions between bio-macromolecules and small molecules by using affinity capillary electrophoresis(ACE), partial filling affinity capillary electrophoresis(FACE), frontal analysis capillary electrophoresis(FACE), and so on. Cell, which plays important role in the absorption, distribution and metabolism of drugs, is the basic unit of life with specific structure and functions. Therefore, it is meaningful to study the affinity between cells and small molecules(or drug candidates). The present study was aimed to develop CE methods for studying the affinity interaction between cells and small molecules, including CZE method for on-line evaluating the anti-platelet aggregation activity of Rhizoma Corydalis(RC) extract, open tubular affinity capillary electrochromatography(OT-ACEC) for evaluating the interaction between platelets and active small molecules, and ACE, PFACE and FACE for studying the interaction between macrophages and small molecules with anti-inflammatory activity. This thesis mainly includes five parts(chapters).The first part is literature review. The significance, objects and methods of affinity interaction study were summarized. Then, the physiological and pathological role of platelet and macrophage were briefly introduced.In the second part, a new CE method was developed for on-line detecting platelet aggregation and evaluating anti-platelet aggregation activity of drugs. Based on the principle of CE in determination of platelet aggregation, the equations used for calculating the anti-platelet aggregation activity of drug were obtained by referring to the turbidimetric method. The anti-platelet aggregation activity of RC extract was evaluated by both CE and turbidimetric method. And then, the differences in sample consumption, values of inhibition rates and the principles of two methods were discussed.In the third part, a platelet-functionalized OT-ACEC method was developed and applied to evaluate the interaction between platelet and small molecules include salvianolic acid B, salvianic acid A sodium, hydroxysafflor yellow A, ferulic acid, chlorogenic acid, sinapic acid and caffeic acid. Besides, the in vitro anti-platelet aggregation(induced by ADP, AA and THR) activity of these molecules was determined by turbidimetric method, and the relationship between anti-platelet aggregation activity of small molecules and their affinity interactions with platelets were discussed.In the fourth part, ACE, PFACE and FACE were applied to evaluate the affinity interaction between macrophage and small molecules. The PFACE was used to evaluate the interaction between macrophages and small molecules qualitatively. Taking advantage of cell tend to be aggregated under electric field, FACE was used to detect the binding constant between macrophage and naringin. Then, macrophage as pseudo stationary phase was used to study the potential active ingredients in lemon extract.The fifth part summarized the innovation points, drawbacks and prospective of the present study. The results showed that CE could be used to on-line detect platelet aggregation and to evaluate anti-platelet aggregation activities of drugs, and to study the affinity interactions between cells and small molecules. In general, as a separation method, CE is a promising method for on-line evaluating interaction between cells and drugs, including determination of binding constant, cell electrophoresis anal ysis, determination of cell secretions, and so on.