Journal of chromatography. A 2014-08-15

Fluorescence derivatization method for sensitive chromatographic determination of zidovudine based on the Huisgen reaction.

Yuki Maeda, Naoya Kishikawa, Kaname Ohyama, Mitsuhiro Wada, Rie Ikeda, Naotaka Kuroda

Index: J. Chromatogr. A. 1355 , 206-10, (2014)

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Abstract

A novel pre-column fluorescence derivatization method for chromatographic analysis of azide compounds was developed based on the Huisgen reaction, which is a specific cycloaddition reaction between an alkyne and an azide. We designed and synthesized a fluorescent alkyne, 2-(4-ethynylphenyl)-4,5-diphenyl-1H-imidazole (DIB-ET) as a reagent. DIB-ET has a lophine skeleton carrying an alkyne acting as fluorophore and reactive center, respectively. In order to evaluate the practicality of DIB-ET, a high-performance liquid chromatography with fluorescence detection method was developed for the determination of zidovudine as a model of azide compound. Zidovudine could be reacted with DIB-ET in the presence of copper(II) sulfate and L-ascorbic acid as catalysts, and the formed derivative was detected fluorometrically. The proposed method allows sensitive and selective determination of zidovudine in plasma samples with the detection limit of 0.28ngmL(-1) at a S/N=3. Finally, the proposed method could be applied to determine the zidovudine concentration in rat plasma after administration of zidovudine without interference from biological components. Crown Copyright © 2014. Published by Elsevier B.V. All rights reserved.