School of Pharmacy, College of Pharmacy, Kaohsiung Medical University, Kaohsiung, Taiwan.
Electrophoresis. 2010 May;31(9):1572-82. doi: 10.1002/elps.200900768.
Phenytoin is a widely used anti-seizure agent, and a good correlation is observed between its concentration in plasma and the clinical effect. We developed a selective CE with UV detection at 200 nm for analysis of free and total levels of phenytoin in human plasma based on MEKC. A sample pretreatment by liquid-liquid extraction with ethyl acetate for determination of total level of phenytoin and serum ultrafiltrate was prepared by ultrafiltration technique (ultrafiltration membrane 30 kDa; 2000 g, 10 min) for determination of free level of phenytoin and subsequent quantification by MEKC was used. MEKC was performed in Tris buffer (25 mM; pH 10.5) containing SDS (180 mM) and EG (13%) as BGE. Hydrodynamic injection for phenytoin determination (0.5 psi 5 s for total level, 2 psi 5 s for free level) was used to introduce samples. The separation voltage was set at 20 kV. Data obtained by MEKC were compared with the results by a validated HPLC method. The MEKC assay of phenytoin exhibited a very good correlation with respect to HPLC by Bland-Altman method. The equations for the Passing-Bablok regression line were as follows: for total level: MEKC=1.0143 x HPLC+0.0976, r(2)=1; for free level: MEKC=1.0013 x HPLC-0.0016, r(2)=1. The proposed method was applied successfully to monitor free and total levels of phentoin in 20 patients with seizures.
苯妥英是一种广泛使用的抗癫痫药物，其在血浆中的浓度与临床效果之间存在良好的相关性。我们基于 MEKC 开发了一种具有 200nm 处紫外检测的选择性 CE，用于分析人血浆中游离和总苯妥英的浓度。通过液-液萃取（乙酸乙酯）预处理样品，用于测定总苯妥英浓度，然后通过超滤技术（超滤膜 30kDa；2000g，10min）制备血清超滤液，用于测定游离苯妥英浓度，并随后通过 MEKC 进行定量。MEKC 在 Tris 缓冲液（25mM；pH10.5）中进行，其中包含 SDS（180mM）和 EG（13%）作为 BGE。对于苯妥英的测定，采用水力注射（总浓度为 0.5psi5s，游离浓度为 2psi5s）进样。分离电压设置为 20kV。通过 MEKC 获得的数据与经过验证的 HPLC 方法的结果进行了比较。通过 Bland-Altman 方法，MEKC 测定苯妥英的结果与 HPLC 非常相关。通过 Passing-Bablok 回归线的方程如下：总浓度：MEKC=1.0143xHPLC+0.0976，r(2)=1；游离浓度：MEKC=1.0013xHPLC-0.0016，r(2)=1。该方法成功应用于 20 例癫痫患者中游离和总苯妥英浓度的监测。
