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Analysis of drug interactions with alpha1-acid glycoprotein using high-performance affinity chromatography
In the circulatory system, interactions between drugs and serum proteins play an important role in determining the pharmacokinetics and pharmacodynamics of drugs. Alpha1-acid glycoprotein (AGP) is an important transport protein involved in this process, as it can bind to numerous basic and neutral drugs in the blood stream. This dissertation examined the use of high-performance affinity chromatography (HPAC) and AGP microcolumns in the analysis of interactions between various drugs and AGP to better understand the transportation of drugs in the circulation. A major portion of this research focused on the development and optimization of a slurry-based entrapment method for preparing AGP microcolumns. The columns that were prepared were found to give entrapped AGP that had good agreement with the binding behavior that is seen for soluble AGP. Application of the entrapped AGP microcolumns was also extended to kinetic studies of multi-site interactions between drugs and AGP by using HPAC and peak profiling. The next portion of this research examined the use of immobilized AGP and ultrafast affinity extraction in free fraction analysis on a two-dimensional affinity system. Conditions needed for the free fraction measurements were examined and optimized. The final system was used to measure the free fractions of several model drugs in serum at typical therapeutic concentrations, and to estimate the binding constants for these drugs with soluble AGP. The use of chromatographic theory and work with model systems was also discussed to provide guidelines for selecting optimal or usable conditions for free fraction analysis by ultrafast affinity extraction for various biological systems. A third portion of this research involved the use of on-line immunoextraction and HPAC to prepare immunoextraction columns containing adsorbed AGP. These columns were used to examine the changes in binding of several drugs to AGP that may occur during the systemic lupus erythematosus (SLE). Both frontal analysis and zonal elution were conducted on the isolated AGP from an SLE patient with a numbers of drugs, with some of these drugs showing significant changes in binding versus that seen with normal AGP. These changes in drug binding to AGP can provide information on how structural changes in AGP during SLE can affect the drug binding properties of this protein.
Bi, Cong, "Analysis of drug interactions with alpha1-acid glycoprotein using high-performance affinity chromatography" (2016). ETD collection for University of Nebraska - Lincoln. AAI10103300.