Róna K, Ary K, Gachályi B, Klebovich I
Department of I. Medicine, Haynal Imre University of Medical Sciences, Budapest, Hungary.
J Chromatogr A. 1996 Apr 12;730(1-2):125-31. doi: 10.1016/0021-9673(95)01227-3.
A sensitive reversed-phase gradient elution high-performance liquid chromatographic method with fluorescence detection has been developed for the determination of alpha-methyldopa (AMD) in human plasma. Separation of the investigated compound and the 3,4-dihydroxyphenylalanine (DOPA) internal standard was achieved on a Nucleosil 7 C18 column with a 5 mM heptanesulphonic acid sodium salt containing 0.05 M potassium dihydrogenphosphate (pH 3.2)-acetonitrile mobile phase. The composition of the mobile phase was changed according to a linear gradient time program. Detection was performed at 270 nm fluorimetric excitation and 320 nm emission. The compounds were isolated from plasma by Bond-Elut C18 solid-phase extraction. The limit of quantitation was found to be 10 ng/ml plasma. The assay was validated with respect to accuracy, precision and system suitability. All validated parameters were found to be within the 20% required limits. On the basis of the sensitivity, linearity and validation parameters the developed analytical method was found to be suitable for application in a bioequivalency study.
已开发出一种灵敏的反相梯度洗脱高效液相色谱法，并采用荧光检测，用于测定人血浆中的α-甲基多巴（AMD）。在所研究的化合物与3,4-二羟基苯丙氨酸（DOPA）内标之间，通过在Nucleosil 7 C18柱上，采用含0.05 M磷酸二氢钾（pH 3.2）的5 mM庚烷磺酸钠盐-乙腈流动相实现分离。流动相的组成根据线性梯度时间程序进行变化。荧光检测在激发波长270 nm和发射波长320 nm下进行。通过Bond-Elut C18固相萃取从血浆中分离出这些化合物。定量限为10 ng/ml血浆。该测定法在准确度、精密度和系统适用性方面进行了验证。所有验证参数均在规定的20%限度内。基于灵敏度、线性和验证参数，所开发的分析方法适用于生物等效性研究。
