Development and validation of a HPAE-PAD method for the quantification of CGP69669A, a sialyl Lewisx mimetic, in skin permeation studies

Abstract

A simple, rapid, precise and specific isocratic HPAE-PAD method for quantification of CGP69669A was developed and validated. CGP69669A is a glycomimetic of sialyl Lewis(x) and an antagonist of E-selectin with potential application in the treatment of inflammatory skin disease. Quantification was performed using a Dionex CarboPac(TM) PA-200 anion-exchange column (3 × 250 mm) with 100 mm NaOH solution as mobile phase, a flow rate of 0.50 mL/min and an injection volume of 10 μL. A quadruple potential waveform was used to detect the carbohydrate (+0.1 V from 0.00 to 0.40 s, -2.0 V from 0.41 to 0.42 s, +0.6 V at 0.43 s and -0.1 V from 0.44 to 0.50 s with current integrated between 0.20 and 0.40 s for detection) and rafinose was employed as an internal standard. The optimized conditions enabled rapid elution of CGP69669A (at 3.0 min) without interference from solvent peaks or substances present in the skin. The method showed good intra- and inter-day precision and accuracy and the response was linear from 1.0 to 25 µg/mL. This is the first validated direct method for the quantification of CGP69669A. It will now be employed in studies investigating the topical and transdermal delivery of CGP69669A in vitro and in vivo and it should also be of use for other applications of this molecule.