LC-MS/MS测定人血浆中依折麦布和总的依折麦布含量及其药动学研究

 目的建立测定人血浆中依折麦布和总的依折麦布浓度的LC-MS/MS方法,进行依折麦布片po给药的药动学研究。方法22名志愿者血样经叔丁基甲醚萃取吹干重组后进样测定依折麦布浓度,另取血样经β-葡萄糖苷酸酶水解后叔丁基醚萃取吹干重组后进样测定总的依折麦布浓度。分析柱:Capcell C₁₈柱(2mm×50mm,5μm),内标(IS)¹³C6-依折麦布。流动相:5mmol·L^-1醋酸铵水溶液(A泵)和乙腈(B泵)组成,梯度洗脱。采用电喷雾去质子化三重四极杆二级串联质谱,多反应监测方式(MRM)测定样品浓度,依折麦布的监测离子对为m/z408.5→m/z270.8,内标为m/z414.5→m/z276.8。结果依折麦布在0.020～20μg·L^-1内线性关系良好,相关系数r=0.9994(n=9)。总的依折麦布在0.25～250μg·L^-1内线性关系良好,相关系数r=0.9998(n=9)。最低定量线(LLOQ)、低、中、高浓度的依折麦布和总的依折麦布质控样品的日内、日间精密度均小于12%,方法回收率在97%～104%内。志愿者口服10mg依折麦布后采血72h,依折麦布和总的依折麦布的药动学参数分别为AUC_0-inf(121.9±41.7)和(536.8±182.7)μg·h·L^-1;AUC_0-t(102.1±31.4)和(478.8±170.0)μg·h·L^-1;ρ_max(4.9±1.6)和(48.46±17.3)μg·L^-1;t_max(5.2±6.4)和(1.5±0.9)h;t_1/2(26.4±28.8)和(22.5±11.0)h;MRT(23.6±3.8)和(19.7±3.8)h。结论该方法符合生物样品分析要求,可用于依折麦布血浆浓度的测定,依折麦布和总的依折麦布的药动学参数与文献相近。

Determination of Ezetimibe and Total Ezetimibe in Human Plasma and Its Pharmacokinetics

OBJECTIVE To develop a LC-MS/MS method for the determination of ezetimibe and total ezetimibe in human plasma and to study its pharmacokinetics in Chinese healthy volunteers.METHODS The analyte and internal standard(¹³C6-ezetimibe)were extracted from plasma samples by liquid-liquid extraction with methyl tert-butyl ether.The reversed-phase chromatographic separation was performed on a Capcell C₁₈ column(2 mm×50 mm,5 μm),and the extract was eluted with a gradient consisting of acetonitrile and 5 mmol·L^-1 ammonium acetate at a flow rate of 0.25 mL·min^-1.The analyte was detected using negative ionization by multiple reaction monitoring mode.The mass transition paires of m/z 408.5→ m/z 207.8 and m/z 408.5→ m/z 207.8 were applied to detect ezetimibe,and ¹³C6-ezetimibe was used to quantify ezetimibe and total ezetimibe.RESULTS A good linearity was obtained in the concentration range of 0.020～20μg·L^-1(r=0.999 4,n=9)for ezetimibe and 0.25～250 μg·L^-1(r=0.999 8,n=9)for total ezetimibe in human plasma.The inter-and intra-day precision(RSD)were less than 12% and the method recoverys were within 97%～104%.The pharmacokinetic parameters of ezetimibe and total ezetimibe after a single oral administration of 10 mg ezetimibe tablets were as follows:AUC_0-inf(121.9±41.7)and(536.8±182.7)μg·h·L^-1;AUC_0-t(102.1±31.4)and(478.8±170.0)μg·h·L^-1;ρ_max(4.9±1.6)and(48.5±17.3)μg·L^-1;t_max(5.2±6.4)and(1.5±0.9)h;t_1/2(26.4±28.8)and(22.5±11.0)h;MRT(23.6±3.8)and(19.7±3.8)h,respectively.CONCLUSION A reliable LC-MS/MS method is developed and validated for the determination of ezetimibe and total ezetimibe in human plasma.The parameters of its pharmacokinetics in Chinese male volunteers are same as reported abroad.