Development and validation of a rapid high-performance liquid chromatography method for the quantification of exenatide.

摘要

The objective was to develop a simple HPLC method to quantify exenatide-a 39 amino acid residue incretin mimetic used in diabetes therapy. To date, only non-validated, sometimes incomplete, gradient methods have been reported in the literature. Isocratic separation was achieved using a C(4) column and a mixed solvent system, A-B-C (48:45:7, v/v/v; pH* 5.2), where A represents KH(2) PO(4) (pH 4.5; 0.1 m) and MeCN (60:40, v/v), B corresponds to NaClO(4) .H(2) O (pH 6.0; 0.2 m) and MeCN (60:40, v/v), and C is water. Exenatide eluted at 3.64 min and the total run time was 6 min. The method was specific and the response was accurate, precise and linear from 0.75 to 25 microg/mL. It was used to quantify exenatide transport across intact and laser-porated porcine skin in vitro as a function of laser fluence [0 (i.e. intact skin), 9 and 15 J/cm(2) , respectively]. Although no permeation was observed using intact skin, cumulative exenatide permeation after 8 h through laser porated skin was 9.6 +/- 6.5 and 12.4 +/- 6.4 microg/cm(2) at fluences of 9 and 15 J/cm(2) , respectively. This is the first validated isocratic method for exenatide quantification and it may be of use in quality control analysis and with other biological matrices. Copyright (c) 2010 John Wiley & Sons, Ltd.