Studies of drug interactions with alpha 1 -acid glycoprotein by using on-line immunoextraction and high-performance affinity chromatography

Abstract

A method that combined on-line immunoextraction with high-performance affinity chromatography was developed to examine the binding of drugs with α1-acid glycoprotein (AGP). Affinity microcolumns containing immobilized polyclonal anti-AGP antibodies were developed that had a capture efficiency of up to 98.4% for AGP and a binding capacity of 0.72 nmol AGP when using a 20 mm × 2.1 mm i.d. microcolumn. These microcolumns were employed in various formats to examine the binding of drugs to normal AGP and AGP that had been adsorbed from serum samples for patients with systemic lupus erythematosus (SLE). Drugs that were screened in zonal elution experiments for their overall binding to these types of AGP included chlorpromazine, disopyramide, imipramine, propranolol, and warfarin. Most of these drugs showed an increase in their binding to the AGP from SLE serum when compared to normal AGP (i.e., an increase of 13–76%); however, disopyramide gave a 21–25% decrease in retention when the same AGP samples were compared. Frontal analysis was used to further evaluate the binding of disopyramide and imipramine to these forms of AGP. Both drugs gave a good fit to a model that involved a combination of saturable and non-saturable interactions with AGP. Changes in the non-saturable interactions accounted for most of variations seen in the binding of disopyramide and imipramine with the AGP samples. The methods used in this study could be adapted for use in personalized medicine and the study of other proteins or drugs using aqueous mixtures or clinical samples.

Thermophoresis for characterizing biomolecular interaction

“…Various analytical techniques that have been used to characterize biomolecular interactions. Some of these techniques include affinity-based separation techniques, which represent the major domain of affinity chromatography, [4][5][6] and affinity capillary electrophoresis [7][8][9] as well as equilibrium dialysis, which is still interesting and widely used especially in drug-protein interaction studies [10][11][12]. On the other hand, biochemical and biophysical techniques are attractive and developed dramatically to be the first choice in studying biomolecular interactions.…”

Analysis of stereoselective drug interactions with serum proteins by high-performance affinity chromatography: A historical perspective

“…This method was then explored for use with HPAC in the mid-1980s for the study of lectin-sugar interactions [39,152,153] and was adapted for use in the study of drug or solute interactions with HSA by the mid-1990s [93,154]. Band-broadening measurements have since been used in various forms to study the binding of drugs and drug metabolites with both HSA and AGP [80,155–157]. …”

“…This difference is determined at one or more flow rates and is used to calculate the dissociation rate constant for the solute with the immobilized binding agent [155]. Peak profiling has been used in recent years to estimate the dissociation rate constants for a number of chiral or achiral drugs and solutes with both HSA and AGP [80,155–158]. …”

“…In addition, detailed thermodynamic studies have been carried out with this protein and such drugs as propranolol, carbamazepine, and warfarin [61,62,76–78]. More recent studies have also employed HPAC and affinity columns to examine the kinetics for some of these interactions [79,80]. …”

“…However, it is now also common to employ more complex models and equations that may involve non-linear fits [7,22]. Examples of such models include those that involve multi-site binding, non-saturable interactions, and combinations of saturable plus non-saturable interactions, such as have been observed for the interactions of some drugs with modified forms of HSA and with AGP or lipoproteins [6,22,76–78,80–84,108–110,132–134]. This approach has further been employed in the study of other systems, such as drug-receptor interactions [135] and the binding of various solutes with lectins or enzymes [136–139].…”