School of Pharmacy, Pacific University, Hillsboro, OR 97123, USA.
J Pharm Biomed Anal. 2009 Dec 5;50(5):987-93. doi: 10.1016/j.jpba.2009.07.004. Epub 2009 Aug 6.
Midazolam (MDZ) is a commonly used benzodiazepine in clinical practice. In addition, its metabolic oxidation is used as a surrogate marker for Cytochrome P450 (CYP) 3A enzyme activity as well. Thus, a new simpler method to measure MDZ and its metabolites is welcomed. Herein we report a new and simple HPLC method with ultraviolet detection for the simultaneous determination of midazolam and its hydroxyl metabolites using lorazepam as an internal standard. A liquid-liquid extraction was used to extract the compounds from rabbit hepatic microsomes and human plasma. The separation was performed on a Zorbax Eclipse XDB C18 column using a mobile phase composed of 0.05 M Na2PO4 (pH 4.5) and acetonitrile mixture (67:33) pumped at 1.2 mL/min. The calibration curves showed good linearity with correlation coefficient higher than 0.999 for all analytes in the range 10-500 ng/mL. Accuracy in the measurement of quality control (QC) samples was in the range 95-106% of the nominal values. The intra-day and inter-day precisions in the measurement of QC samples were less than 11% coefficient of variation. Although less sensitive than GC-MS, the proposed method was adequately sensitive to measure midazolam hydroxylase activity as a marker for CYP3A activity, and was applied to measure midazolam pharmacokinetics in human plasma.
咪达唑仑（MDZ）是临床实践中常用的苯二氮䓬类药物。此外，其代谢氧化也被用作细胞色素 P450（CYP）3A 酶活性的替代标志物。因此，人们欢迎一种新的更简单的方法来测量 MDZ 及其代谢物。在此，我们报告了一种新的简单 HPLC 方法，该方法采用紫外检测，使用劳拉西泮作为内标，同时测定兔肝微粒体和人血浆中的咪达唑仑及其羟基代谢物。采用液-液萃取法从兔肝微粒体和人血浆中提取化合物。分离在 Zorbax Eclipse XDB C18 柱上进行，流动相由 0.05 M Na2PO4（pH 4.5）和乙腈混合物（67:33）组成，以 1.2 mL/min 的速度泵入。校准曲线在 10-500 ng/mL 范围内，所有分析物的相关系数均高于 0.999，呈良好线性。在 QC 样品测量中的准确度在标称值的 95-106%范围内。QC 样品测量的日内和日间精密度变异系数均小于 11%。虽然灵敏度不如 GC-MS 高，但该方法足够灵敏，可用于测量咪达唑仑羟化酶活性作为 CYP3A 活性的标志物，并应用于测量人血浆中咪达唑仑的药代动力学。
