title: Validation of a method for the quantitation of ghrelin and unacylated ghrelin by HPLC.
abstract: An HPLC/UV method was first optimized for the separation and quantitation of human acylated and unacylated (or des-acyl) ghrelin from aqueous solutions. This method was validated by an original approach using accuracy profiles based on tolerance intervals for the total error measurement. The concentration range that achieved adequate accuracy extended from 1.85 to 59.30microM and 1.93 to 61.60microM for acylated and unacylated ghrelin, respectively. Then, optimal temperature, pH and buffer for sample storage were determined. Unacylated ghrelin was found to be stable in all conditions tested. At 37 degrees C acylated ghrelin was stable at pH 4 but unstable at pH 7.4, the main degradation product was unacylated ghrelin. Finally, this validated HPLC/UV method was used to evaluate the binding of acylated and unacylated ghrelin to liposomes.
authors.affiliation: Université catholique de Louvain, Louvain Drug Research Institute, Unité de Pharmacie Galénique, UCL 73.20, Avenue Emmanuel Mounier 73, 1200 Brussels, Belgium.
authors.forename: Edith
authors.initials: E
authors.lastname: Staes
authors.forename: Eric
authors.initials: E
authors.lastname: Rozet
authors.forename: Bernard
authors.initials: B
authors.lastname: Ucakar
authors.forename: Philippe
authors.initials: P
authors.lastname: Hubert
authors.forename: Véronique
authors.initials: V
authors.lastname: Préat
mesh_terms: D000215:Acylation; D002626:Chemistry, Pharmaceutical / Q000379:methods / Q000592:standards*; D002851:Chromatography, High Pressure Liquid / Q000379:methods / Q000592:standards; D054439:Ghrelin / Q000032:analysis*; D006801:Humans
chemical_list: D054439:Ghrelin