Methods of Determining Plasma and Tissue Binding of Drugs

Abstract

The available techniques for the investigation of drug binding to plasma and tissues protein are reviewed and the advantages and disadvantages of the various techniques stated. A comparison of different plasma protein binding techniques is made which shows that the size of the unbound fraction of drug may be influenced by the method used. Protein binding may be assayed by methods including equilibrium dialysis, ultrafiltration, ultracentrifugation, gel filtration, binding to albumin microspheres and circular dichroism. Tissue binding techniques can involve testing binding to isolated organs, tissue slices, homogenates and isolated subcellular particles. Details of the available methods to compute pharmacokinetic constants are given. Stereoselective binding has been investigated for a limited number of drugs and the difference in the binding of 2 enantiomers is usually modest. The measurement of the binding constants is often required to characterise the drug-protein interaction. Mathematical and graphical methods to compute the pharmacokinetic parameters are discussed. The implications of binding on the volume of distribution and clearance of drugs are examined.

In vitro blood distribution and plasma protein binding of the tyrosine kinase inhibitor imatinib and its active metabolite, CGP74588, in rat, mouse, dog, monkey, healthy humans and patients with acute lymphatic leukaemia

“…Differences in plasma protein binding of a drug between species or individuals have an impact on pharmacokinetics and can influence the pharmacological activity [12,13]. For imatinib elevated levels of AGP have been related to in vivo resistance in mouse [9] and humans [14,15], whereas other investigators did not observe any relevant binding of imatinib to AGP employing fluorescence quenching [16].…”

Preclinical Species and Human Disposition of PF-04971729, a Selective Inhibitor of the Sodium-Dependent Glucose Cotransporter 2 and Clinical Candidate for the Treatment of Type 2 Diabetes Mellitus

“…The binding of PF-04971729 to rat, dog, and human plasma proteins was determined using the equilibrium dialysis method (Pacifici and Viani, 1992) using cellulose membranes (HTDialysis, Gales Ferry, CT) with a molecular mass cutoff of 12 to 14 kDa. For plasma protein binding studies, plasma was fortified with PF-04971729 at concentrations of 2.3 and 23 M, which reflect a theoretical plasma concentration anticipated in humans and a multiple thereof to assess saturation of plasma protein binding.…”

Effect of Albumin on Phenytoin and Tolbutamide Metabolism in Human Liver Microsomes: An Impact More Than Protein Binding

“…For example, it has been noted that during one passage through the brain, in many cases, more drug than "free drug" can penetrate through the blood-brain barrier (Spector, 2000). Similarly, the hepatic uptake of many lipophilic compounds is not necessarily restricted by protein binding (Kurz and Fichtl, 1983;Pacifici and Viani, 1992), and it has been found that the antifungal activity of itraconazole and ketoconazole is not diminished despite extensive binding albumin (Schäfer-Korting et al, 1995). In agreement, Tran et al (1997) have reported that the K i value of stiripentol obtained in microsomal studies is more consistent with total plasma concentration than unbound concentration.…”