Malli Danai, Gikas Evagelos, Vavagiannis Andreas, Kazanis Michael, Daniilides Konstantinos, Gennimata Dimitra, Panderi Irene
University of Athens, School of Pharmacy, Division of Pharmaceutical Chemistry, Panepistimiopolis, Zografou, GR-157 71 Athens, Greece.
Anal Chim Acta. 2007 Sep 5;599(1):143-50. doi: 10.1016/j.aca.2007.08.004. Epub 2007 Aug 6.
A sensitive and selective high-performance liquid chromatographic method has been developed and validated for the determination of nateglinide in human plasma. Nateglinide and the internal standard, undecylenic acid, were extracted from plasma by liquid-liquid extraction using a mixture of ethyl acetate-diethyl ether, 50:50 (v/v). Pre-column derivatization reaction was performed using a coumarin-type fluorescent reagent, N-(7-methoxy-4-methyl-2-oxo-2H-6-chromenyl)-2-bromoacetamide. The derivatization proceeded in acetone in the presence of potassium carbonate and catalyzed by 18-crown-6 ether. The fluorescent derivatives were separated under isocratic conditions on a Hypersil BDS-C8 analytical column (250.0 mm x 2.1 mm i.d., particle size 5 microm) with a mobile phase that consisted of 65% acetonitrile in water and pumped at a flow rate of 0.50 mL min(-1). The excitation and emission wavelengths were set at 345 and 435 nm, respectively. The assay was linear over a concentration range of 0.05-16.00 microg mL(-1) for nateglinide with a limit of quantitation of 0.05 microg mL(-1). Quality control samples (0.05, 4.50 and 16.00 microg mL(-1)) in five replicates from five different runs of analysis demonstrated intra-assay precision (%coefficient of variation <6.8%), inter-assay precision (%coefficient of variation <1.6%) and an overall accuracy (%relative error) less than -3.4%. The method can be used to quantify nateglinide in human plasma covering a variety of pharmacokinetic or bioequivalence studies.
已开发并验证了一种灵敏且具选择性的高效液相色谱法，用于测定人血浆中的那格列奈。采用乙酸乙酯 - 乙醚（50:50，v/v）混合液通过液 - 液萃取从血浆中提取那格列奈和内标十一烯酸。使用香豆素型荧光试剂N - (7 - 甲氧基 - 4 - 甲基 - 2 - 氧代 - 2H - 6 - 苯并二氢吡喃 - 2 - 基) - 2 - 溴乙酰胺进行柱前衍生化反应。衍生化反应在丙酮中于碳酸钾存在下进行，并由18 - 冠 - 6醚催化。荧光衍生物在等度条件下于Hypersil BDS - C8分析柱（250.0 mm×2.1 mm内径，粒径5微米）上分离，流动相为65%乙腈的水溶液，流速为0.50 mL min⁻¹。激发波长和发射波长分别设定为345和435 nm。那格列奈在0.05 - 16.00 μg mL⁻¹浓度范围内呈线性，定量限为0.05 μg mL⁻¹。来自五次不同分析运行的五个重复的质量控制样品（0.05、4.50和16.00 μg mL⁻¹）显示批内精密度（变异系数%<6.8%）、批间精密度（变异系数%<1.6%）以及总体准确度（相对误差%）小于 - 3.4%。该方法可用于定量人血浆中的那格列奈，适用于各种药代动力学或生物等效性研究。
