1. J Pharm Biomed Anal. 2010 Feb 5;51(3):633-9. Epub 2009 Oct 2.

Validation of a method for the quantitation of ghrelin and unacylated ghrelin by 
HPLC.

Staes E, Rozet E, Ucakar B, Hubert P, Préat V.

Université catholique de Louvain, Louvain Drug Research Institute, Unité de
Pharmacie Galénique, UCL 73.20, Avenue Emmanuel Mounier 73, 1200 Brussels,
Belgium.

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.

PMID: 19875264 [PubMed - in process]