Characterization of the protein binding of chiral drugs by high-performance affinity chromatography. Interactions of R- and S-ibuprofen with human serum albumin.
Zonal elution and high-performance affinity chromatography were used to study the different binding characteristics of R- and S-ibuprofen with the protein human serum albumin (HSA). This was done by injecting small amounts of R- and S-ibuprofen onto an immobilized HSA column in the presence of a mobile phase that contained a known concentration of R- or S-ibuprofen as a competing agent. These studies indicated that R- and S-ibuprofen had one common binding site on the immobilized HSA column. In addition, S-ibuprofen had at least one other major binding region. The association equilibrium constant for R-ibuprofen with HSA was found to be 5.3 x 10(5) M-1 at pH 6.9 and 25 degrees C. Under the same conditions, the association constants for S-ibuprofen at its two sites were 1.1 x 10(5) M-1 and 1.2 x 10(5) M-1. The S-ibuprofen sites were present in about a 1:1 ratio and appeared to exhibit some allosteric interactions at high S-ibuprofen concentrations. The chromatographic technique used in this work is a general one which can be adapted for use in studying the interactions of other chiral compounds with either HSA or additional proteins.
Hage
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DS
Department of Chemistry, University of Nebraska, Lincoln 68588-0304.
Noctor
T A
TA
Wainer
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IW
D001667:Binding, Competitive; D002846:Chromatography, Affinity / Q000379:methods*; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007052:Ibuprofen / Q000737:chemistry / Q000378:metabolism*; D008956:Models, Chemical; D012709:Serum Albumin / Q000378:metabolism*; D013237:Stereoisomerism
D012709:Serum Albumin; D007052:Ibuprofen