Chromatographic analysis of carbamazepine binding to human serum albumin

Hee Seung Kim, David S Hage

Research output: Contribution to journalArticle

75 Citations (Scopus)

Abstract

In this study, high-performance affinity chromatography was used to characterize the binding of carbamazepine to an immobilized human serum albumin (HSA) column. Frontal analysis was first used to determine the association equilibrium constant and binding capacity for carbamazepine on this column at various temperatures. The non-specific binding of carbamazepine within the column was also considered. The results indicated that carbamazepine had a single binding site on HSA with an association equilibrium constant of 5.3 × 103 M-1 at pH 7.4 and 37°C. This was confirmed through zonal elution self-competition studies. The value of ΔG for this reaction was -5.35 kcal/mol at 37°C, with an associated change in enthalpy (ΔH) of -6.45 kcal/mol and a change in entropy (ΔS) of -3.56 cal/mol K. The location of this binding region was examined by competitive zonal elution experiments using probe compounds with known sites on HSA. It was found that carbamazepine had direct competition with l-tryptophan, a probe for the indole-benzodiazepine site of HSA, but allosteric interactions with probes for the warfarin, tamoxifen and digitoxin sites. Changes in the pH, ionic strength, and organic modifier content of the mobile phase were used to identify the predominant forces in the carbamazepine-HSA interaction.

Original languageEnglish (US)
Pages (from-to)57-66
Number of pages10
JournalJournal of Chromatography B: Analytical Technologies in the Biomedical and Life Sciences
Volume816
Issue number1-2
DOIs
StatePublished - Feb 25 2005

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Chromatographic analysis
Carbamazepine
Serum Albumin
Chromatography
Equilibrium constants
Association reactions
Digitoxin
Affinity chromatography
Entropy
Warfarin
Tamoxifen
Ionic strength
Benzodiazepines
Affinity Chromatography
Tryptophan
Osmolar Concentration
Enthalpy
Binding Sites
Temperature

Keywords

  • Carbamazepine
  • Frontal analysis
  • HSA
  • High-performance affinity chromatography
  • Zonal elution

ASJC Scopus subject areas

  • Analytical Chemistry
  • Biochemistry
  • Clinical Biochemistry
  • Cell Biology

Cite this

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title = "Chromatographic analysis of carbamazepine binding to human serum albumin",
abstract = "In this study, high-performance affinity chromatography was used to characterize the binding of carbamazepine to an immobilized human serum albumin (HSA) column. Frontal analysis was first used to determine the association equilibrium constant and binding capacity for carbamazepine on this column at various temperatures. The non-specific binding of carbamazepine within the column was also considered. The results indicated that carbamazepine had a single binding site on HSA with an association equilibrium constant of 5.3 × 103 M-1 at pH 7.4 and 37°C. This was confirmed through zonal elution self-competition studies. The value of ΔG for this reaction was -5.35 kcal/mol at 37°C, with an associated change in enthalpy (ΔH) of -6.45 kcal/mol and a change in entropy (ΔS) of -3.56 cal/mol K. The location of this binding region was examined by competitive zonal elution experiments using probe compounds with known sites on HSA. It was found that carbamazepine had direct competition with l-tryptophan, a probe for the indole-benzodiazepine site of HSA, but allosteric interactions with probes for the warfarin, tamoxifen and digitoxin sites. Changes in the pH, ionic strength, and organic modifier content of the mobile phase were used to identify the predominant forces in the carbamazepine-HSA interaction.",
keywords = "Carbamazepine, Frontal analysis, HSA, High-performance affinity chromatography, Zonal elution",
author = "Kim, {Hee Seung} and Hage, {David S}",
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AU - Hage, David S

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N2 - In this study, high-performance affinity chromatography was used to characterize the binding of carbamazepine to an immobilized human serum albumin (HSA) column. Frontal analysis was first used to determine the association equilibrium constant and binding capacity for carbamazepine on this column at various temperatures. The non-specific binding of carbamazepine within the column was also considered. The results indicated that carbamazepine had a single binding site on HSA with an association equilibrium constant of 5.3 × 103 M-1 at pH 7.4 and 37°C. This was confirmed through zonal elution self-competition studies. The value of ΔG for this reaction was -5.35 kcal/mol at 37°C, with an associated change in enthalpy (ΔH) of -6.45 kcal/mol and a change in entropy (ΔS) of -3.56 cal/mol K. The location of this binding region was examined by competitive zonal elution experiments using probe compounds with known sites on HSA. It was found that carbamazepine had direct competition with l-tryptophan, a probe for the indole-benzodiazepine site of HSA, but allosteric interactions with probes for the warfarin, tamoxifen and digitoxin sites. Changes in the pH, ionic strength, and organic modifier content of the mobile phase were used to identify the predominant forces in the carbamazepine-HSA interaction.

AB - In this study, high-performance affinity chromatography was used to characterize the binding of carbamazepine to an immobilized human serum albumin (HSA) column. Frontal analysis was first used to determine the association equilibrium constant and binding capacity for carbamazepine on this column at various temperatures. The non-specific binding of carbamazepine within the column was also considered. The results indicated that carbamazepine had a single binding site on HSA with an association equilibrium constant of 5.3 × 103 M-1 at pH 7.4 and 37°C. This was confirmed through zonal elution self-competition studies. The value of ΔG for this reaction was -5.35 kcal/mol at 37°C, with an associated change in enthalpy (ΔH) of -6.45 kcal/mol and a change in entropy (ΔS) of -3.56 cal/mol K. The location of this binding region was examined by competitive zonal elution experiments using probe compounds with known sites on HSA. It was found that carbamazepine had direct competition with l-tryptophan, a probe for the indole-benzodiazepine site of HSA, but allosteric interactions with probes for the warfarin, tamoxifen and digitoxin sites. Changes in the pH, ionic strength, and organic modifier content of the mobile phase were used to identify the predominant forces in the carbamazepine-HSA interaction.

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