de Sousa Maia Maria Bernadete, do Nascimento Demétrius Fernandes, Martins Ismael Leite, Cunha Adriano Nunes, de Lima Francisco Evanir Gonçalves, Bezerra Fernando Antônio Frota, de Moraes Manoel Odorico, de Moraes Maria Elisabete Amaral
Department of Physiology and Pharmacology, Federal University of Pernambuco, Av. Prof. Moraes Rêgo S/N, Cidade Universitária, 50670-901 Recife, Pernambuco, Brazil.
J Pharm Biomed Anal. 2007 Oct 18;45(2):304-11. doi: 10.1016/j.jpba.2007.08.001. Epub 2007 Aug 9.
A fast and sensitive method to quantify oxcarbazepine (OXC) and its active metabolite, 10,11-dihydro-10-hydroxycarbamazepine (MHD) in human plasma using HPLC-MS/MS has been developed. The method involved liquid-liquid extraction (LLE), with diethyl ether-diclhoromethane (60:40v/v) using deuterade carbamazepine (d10-carbamazepine) as internal standard (IS). The analytes and IS were separated using an isocratic mobile phase (acetonitrile/water (50:50v/v)+20 mM acetic acid) on the analytical column Phenomenex Luna C18 5 microm (150 mm x 4.6 mm) at room temperature. Detection was performed by a Micromass Quatro LC mass spectrometer in the reaction monitoring mode using positive electrospray ionization (ESI+). The MS-MS ion transition monitored were m/z 253>208 for OXC, m/z 255>194 for MHD and m/z 247>204 for IS. Over the range 20-5250 ng/ml for OXC and 40-10,500 ng/ml for MHD, the calibration curves were defined by the following equations: y = 0.00568 + 0.00296x -5.70e - 8x(2) and y = 0.00749 + 0.00178x - 5.70e - 8x(2) for OXC and MHD, respectively. All coefficient of determination (r(2)) were close to unity (0.9986-0.9994). The lower limits of quantification obtained as a result of the LLE procedure was 20 ng/ml for OXC and 40 ng/ml for MHD. The statistical evaluation of the developed method was conducted by examining within-batch and between-batch precision data, which were within the required limits. The suitability of the assay for pharmacokinetics studies was determined by measuring OXC and MHD concentration after administration of a single 10 ml of OXC oral suspension (6%) in plasma human of healthy volunteers.
已开发出一种快速灵敏的方法，使用高效液相色谱-串联质谱法（HPLC-MS/MS）定量测定人血浆中的奥卡西平（OXC）及其活性代谢物10,11-二氢-10-羟基卡马西平（MHD）。该方法采用液-液萃取（LLE），以氘代卡马西平（d10-卡马西平）作为内标（IS），使用乙醚-二氯甲烷（60:40 v/v）。在室温下，使用等度流动相（乙腈/水（50:50 v/v）+20 mM乙酸）在Phenomenex Luna C18 5微米（150 mm×4.6 mm）分析柱上分离分析物和内标。通过Micromass Quatro LC质谱仪在反应监测模式下使用正电喷雾电离（ESI+）进行检测。监测的MS-MS离子跃迁为：OXC的m/z 253>208，MHD的m/z 255>194，内标的m/z 247>204。在OXC浓度范围为20-5250 ng/ml和MHD浓度范围为40-10500 ng/ml时，校准曲线分别由以下方程定义：OXC为y = 0.00568 + 0.00296x -5.70e - 8x(2)，MHD为y = 0.00749 + 0.00178x - 5.70e - 8x(2)。所有决定系数（r(2)）均接近1（0.9986-0.9994）。液-液萃取程序得到的定量下限为：OXC为20 ng/ml，MHD为40 ng/ml。通过检查批内和批间精密度数据对所开发方法进行统计评估，这些数据在规定范围内。通过测量健康志愿者口服10 ml单剂量奥卡西平口服混悬液（6%）后血浆中OXC和MHD的浓度，确定该测定法对药代动力学研究的适用性。
