大鼠体内咪达唑仑及其代谢物1’-羟基咪达唑仑的药动学研究

目的建立快速、灵敏的LC-MS/MS法测定大鼠血浆中咪达唑仑及其代谢物1’-羟基咪达唑仑的血药浓度,并研究其在大鼠体内的药动学行为。方法按1 kg体质量给予大鼠20 mg.kg-1咪达唑仑的剂量灌胃给药,LC-MS/MS法测定不同时间点的血药浓度。色谱条件:色谱柱为Alltima C18(150 mm×2.1mm,5μm),流动相为甲醇-0.025%(体积分数)甲酸(体积比85∶15),流速为0.3 mL.min-1,柱温为40℃;质谱条件:采用正离子模式,以多反应离子监测(MRM)配对离子方式进行定量。采用PKSolution2.0软件计算其药动学参数。结果咪达唑仑及其代谢物1’-羟基咪达唑仑的线性范围分别为1～1 000ng.mL-1和1～100 ng.mL-1,最低检测限分别为0.5、0.25 ng.mL-1,专属性、精密度、稳定性和基质效应都满足要求。咪达唑仑、1’-羟基咪达唑仑的t1/2分别为(0.748±0.05)h、(1.65±0.33)h;AUC分别为(1 038.3±413.2)ng.mL-1.h、(169.5±38.6)ng.mL-1.h。结论本方法专属性强、灵敏度高、准确性好,可用于咪达唑仑及其代谢物1’-羟基咪达唑仑的含量测定及药动学研究。

LC-MS/MS determination of midazolam and 1'-hydroxymidazolam, its main metabolite in rat plasma and its pharmacokinetics

Objective To establish an LC-MS/MS method for the determination of midazolam and 1′-hydroxymidazolam,its main metabolite in rat plasma and study its pharmacokinetics.Methods Midazolam and 1′-hydroxymidazolam were determined by LC-MS/MS after oral administration of 20 mg·kg-1 of midazolam to rats.Midazolam and 1′-hydroxy midazolam were separated by Alltima C18 column(150 mm×2.1 mm,5 μm) with the mobile phase of methanol and 0.025% formic acid solution(85∶15) at a flow rate of 0.3 mL·min-1.The temperature of column was 40 ℃.The LC-MS/MS system was operated using an electrospray ionization probe in the positive ion mode with multiple reaction ion monitoring(MRM).The pharmacokinetic parameters were calculated by PKSolution software(2.0 version).Results The linear range of midazolam and 1′-hydroxymidazolam were 1～1 000 ng·mL-1 and 1～100 ng·mL-1,respectively.The lowest limit of quantification(LOQ) of midazolam and 1′-hydroxymidazolam were 0.5 ng·mL-1 and 0.25 ng·mL-1(S/N>3),respectively.The accuracy,precision,stability and matrix effect were qualified.The plasma elimination half-life time(t1/2) of midazolam and 1′-hydroxymidazolam were(0.748±0.05)h and(1.65±0.33) h,respectively.The area under the curves(AUC) were(1 038.3±413.2) ng·mL-1·h and(169.5±38.6) ng·mL-1·h,respectively.Conclusion A sensitive,accurate and suitable LC-MS/MS method for the determination midazolam and 1′-hydroxymidazolam was developed and successfully applied for the pharmacokinetic study of midazolam and 1′-hydroxymidazolam,its main metabolite in rats.