Impact of pH on Plasma Protein Binding in Equilibrium Dialysis

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• 作者：Christopher J. Kochansky ; Daniel R. McMasters ; Ping Lu ; Kenneth A. Koeplinger ; Haley H. Kerr ; Magang Shou ; Kenneth R. Korzekwa
• 刊名：Molecular Pharmaceutics
• 出版年：2008
• 出版时间：June 2008
• 年：2008
• 卷：5
• 期：3
• 页码：438 - 448
• 全文大小：1021K
• 年卷期：v.5,no.3(June 2008)
• ISSN：1543-8392

文摘

Many pharmacokinetic analyses require unbound plasma concentrations, including prediction of clearance, volume of distribution, drug−drug interactions, brain uptake analysis, etc. It is most often more convenient to measure the total drug concentration in plasma rather than the unbound drug concentration. To arrive at unbound plasma concentrations, separate in vitro determinations of the plasma protein binding of a drug are usually carried out in serum or in plasma, and the plasma pharmacokinetic results are then mathematically adjusted by this fraction unbound (f_u,p). Plasma protein binding or the drug fraction unbound in plasma (f_u,p) is known to be affected by protein, drug, free fatty acid concentrations, lipoprotein partitioning, temperature, pH, and the presence or absence of other drugs/displacing agents within plasma samples. Errors in f_u,p determination caused by lack of adequate pH control in newer assay formats for plasma protein binding (e.g., 96-well equilibrium thin walled polypropylene dialysis plates) will have significant drug-specific impact on these pharmacokinetic calculations. Using a diverse set of 55 drugs and a 96-well equilibrium dialysis plate format, the effect of variable pH during equilibrium dialysis experiments on measured values of f_u,p was examined. Equilibrium dialysis of human plasma against Dulbecco’s phosphate buffered saline at 37 °C under an air or 10% CO₂ atmosphere for 22 h resulted in a final pH of approximately 8.7 and 7.4, respectively. The ratio of f_u,p at pH 7.4 (10% CO₂) vs pH 8.7 (air) was ≥2.0 for 40% of the 55 compounds tested. Only one of the 55 compounds tested had a ratio <0.9. Select compounds were further examined in rat and dog plasma. In addition, physicochemical properties were calculated for all compounds using ACD/Labs software or Merck in-house software and compared to plasma protein binding results. Changes in plasma protein binding due to pH increases which occurred during the equilibrium dialysis experiment were not species specific but were drug-specific, though nonpolar, cationic compounds had a higher likely hood of displaying pH-dependent binding. These studies underscore the importance of effectively controlling pH in plasma protein binding studies.