LC-MS/MS法测定人血浆中替加色罗的浓度及其在生物等效性研究中的应用

Determination of tegaserod in human plasma by liquid chromatography-tandem mass spectrometry and its application to bioequivalence study

目的：建立灵敏、快速的LC—MS／MS法测定人血浆中替加色罗，并用于制剂生物等效性研究。方法：血浆样品经有机溶剂提取后，以乙腈-5mmol·L^-1乙酸铵-甲酸（70：30：0．01）为流动相，采用Zorbax XDB—C18柱（150mm×4．6mm，5μm）分离，通过电喷雾离子化四极杆串联质谱，以多反应监测（MRM）方式进行检测。用于定量分析的离子反应分别为m／z302→173（替加色罗）和m／z256→167（内标苯海拉明）。结果：替加色罗测定方法的线性范围为0．010—10ng·mL^-1，定量下限为0．010ng·mL^-1，日内、日间精密度（RSD）均〈7．3％，准确度（RE）在±1．4％之内。应用此法研究比较了22例健康受试者单剂量口服替加色罗参比制剂和受试制剂6mg后的主要药动学参数，受试制剂和参比制剂Tmax分别为（0．86±0．22）和（1．01±0．24）h，Cmax分别为（2．21±0．69）和（2．05±0．64）ng·mL^-1，t1/2α分别为（1．18±0．44）和（1．24±0．56）h，t1/2β分别为（10．10±3．07）和（8．81±2．35）h，AUC0-36h分别为（6．35±2．48）和（6．47±1．99）ng·h·mL^-1，AUC0-∞。分别为（6．69±2．59）和（6．70±2．03）ng·h·mL^1。马来酸替加色罗分散片的相对生物利用度平均为（98．2±22．1）％。结论：该法灵敏、快速、准确，适用于替加色罗制剂的人体生物等效性评价。

Objective：To develop and validate a liquid chromatographic-tandem mass spectrometric（LC-MS/MS） method for the determination of tegaserod in human plasma. Methods：Tegaserod and internal standard, diphenhydramine were isolated from plasma by liquid-liquid extraction. The chromatographic separation was achieved within 3 min on Zorbax XDB-C18 column（ 150 mm ×4.6 mm,5μm）with an isocratic mobile phase consisted of acetonitrile-5 mmol· L^- 1 ammonium acetate-formic acid （ 70 ： 30 ： 0.01 ）at the rate of 0.5 mL·min^-1 Multiple reaction monitoring（MRM） transitions of 302→173 and m/z 256→167 were measured in positive mode for tegaserod and the internal standard, respectively. Results： The linear concentration range of the calibration curve for tegaserod was 0. 010 -10 ng·mL^-1. The lower limit of quantification was 0. 010 ng·mL^-1 The intra- and inter-clay relative standard deviations （RSD） across three validation runs over the entire concentration range were less than 7.3% , the accuracies （RE）were within ± 1.4%. After an oral close of 6 mg, the pharmacokinetic parameters of tegaserod in test and reference tablets were as follows ： Tmax （0.86 ± 0.22） and （ 1.01 ± 0.24 ） h, Cmax （ 2. 21 ± 0.69 ） and （ 2. 05 ± 0. 64 ） ng · mL^-1, t1/2 α （1. 18 ±0.44）and（1.24 ±0.56）h, t1/2 β（10. 10 ±3.07）and （8.81 ±2.35）h, AUC0-36h（6.35 ±2.48）and（6.47 ± 1.99） ng ·h· mL^-1 , AUC0-∞ （6.69 ±2.59） and （6.70 ± 2.03） ng ·h· mL^-1, respectively. The relative bioavailability of tegaserod dispersible tablet was （98.2 ± 22.1 ） %. Conclusion ： The method was sensitive, rapid, accurate and suitable for bioequivalence study of tegaserod in human subjects.