Chromatographic analysis of acetohexamide binding to glycated human serum albumin.
Acetohexamide is a drug used to treat type II diabetes and is tightly bound to the protein human serum albumin (HSA) in the circulation. It has been proposed that the binding of some drugs with HSA can be affected by the non-enzymatic glycation of this protein. This study used high-performance affinity chromatography to examine the changes in acetohexamide-HSA binding that take place as the glycation of HSA is increased. It was found in frontal analysis experiments that the binding of acetohexamide to glycated HSA could be described by a two-site model involving both strong and weak affinity interactions. The average association equilibrium constant (K(a)) for the high affinity interactions was in the range of 1.2-2.0×10(5)M(-1) and increased in moving from normal HSA to HSA with glycation levels that might be found in advanced diabetes. It was found through competition studies that acetohexamide was binding at both Sudlow sites I and II on the glycated HSA. The K(a) for acetohexamide at Sudlow site I increased by 40% in going from normal HSA to minimally glycated HSA but then decreased back to near-normal values in going to more highly glycated HSA. At Sudlow site II, the K(a) for acetohexamide first decreased by about 40% and then increased in going from normal HSA to minimally glycated HSA and more highly glycated HSA. This information demonstrates the importance of conducting both frontal analysis and site-specific binding studies in examining the effects of glycation on the interactions of a drug with HSA.
Joseph
K S
KS
Chemistry Department, University of Nebraska, Lincoln, NE 68588-0304, USA.
Anguizola
Jeanethe
J
Jackson
Abby J
AJ
Hage
David S
DS
D000092:Acetohexamide / Q000737:chemistry* / Q000378:metabolism; D002846:Chromatography, Affinity / Q000379:methods*; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D003920:Diabetes Mellitus / Q000188:drug therapy / Q000378:metabolism; D017127:Glycation End Products, Advanced; D006801:Humans; D007004:Hypoglycemic Agents / Q000737:chemistry / Q000378:metabolism; D011485:Protein Binding; D012044:Regression Analysis; D012709:Serum Albumin / Q000737:chemistry* / Q000378:metabolism; D014364:Tryptophan / Q000737:chemistry / Q000378:metabolism; D014859:Warfarin / Q000737:chemistry / Q000378:metabolism; D000093423:Glycated Serum Albumin
D017127:Glycation End Products, Advanced; D007004:Hypoglycemic Agents; D012709:Serum Albumin; D014859:Warfarin; D014364:Tryptophan; D000092:Acetohexamide; D000093423:Glycated Serum Albumin