LC-MS/MS法测定大鼠血浆伏立康唑、伏立康唑氮-氧化物浓度及药动学应用研究

Determination of Voriconazole and Voriconazole N-oxide in Rat Plasma by LC-MS/MS and Pharmacokinetic Application Study

目的:建立液相色谱-串联质谱(LC-MS/MS)法测定大鼠血浆中伏立康唑及伏立康唑氮-氧化物浓度,并探索灌胃给予伏立康唑后两者在大鼠体内的药动学过程。方法:样品预处理选用甲基叔丁基醚作萃取剂,通过液-液萃取法消除基质效应。采用Welch Ultimate UHPLC XB-C_(18)(50 mm×2.1 mm;1.8μm)色谱柱;以乙腈-水(含2 mmol·L^(-1)甲酸铵,0.1%甲酸)为流动相进行梯度洗脱;柱温:35℃;流速:0.35 ml·min^(-1);分析时长:4 min;质谱方法应用电喷雾离子源,多反应监测模式进行正离子检测。利用该方法分别测定大鼠灌胃给予66.7 mg·kg^(-1)伏立康唑后各时间点伏立康唑及伏立康唑氮-氧化物的血药浓度,分析其在大鼠体内的药动学过程。结果:伏立康唑及伏立康唑氮-氧化物在0.5~500 ng·ml^(-1)范围内线性关系良好(r>0.9990),定量下限均为0.5 ng·ml^(-1)。用建立的方法测定伏立康唑及伏立康唑氮-氧化物血药浓度,拟合得到其药动学参数:药时曲线下面积(AUC)_(0-t)分别为(54.66±20.30)μg·ml^(-1)·h,(10.88±5.30)μg·ml^(-1)·h;AUC_(0-∞)分别为(54.66±20.30)μg·ml^(-1)·h,(13.50±2.10)μg·ml^(-1)·h;清除率(CL)分别为(4.19±0.90)L·min^(-1)·kg^(-1),(18.98±3.70)L·min^(-1)·kg^(-1);药峰浓度(C_(max))分别为(19.14±11.90)μg·ml^(-1),(0.95±0.20)μg·ml^(-1);半衰期(t_(1/2))分别为(5.89±6.10)h,(6.11±3.20)h。结论:该方法快速、简便、准确度高且重复性好,可用于伏立康唑及伏立康唑氮-氧化代谢产物在大鼠体内的药动学研究。

Objective:To establish a method of liquid chromatography-mass spectrometry(LC-MS/MS)for the simultaneous determination of voriconazole and voriconazole N-oxide in rat plasma,and to explore their pharmacokinetic process in rats after voriconazole administration by gavage.Methods:Methyl tertiary butyl ether was selected as the extraction agent and liquid-liquid extraction method was applied to eliminating the influence of matrix effect on sample detection.The separation was achieved on a Welch Ultimate UHPLC XB-C_(18)(50 mm×2.1 mm;1.8μm)column with gradient elution of acetonitrile/water(containing 2 mmol·L^(-1) ammonium formate and 0.1%formic acid)at a flow rate of 0.35 ml·min~1,and the column temperature was 35℃.Analysis duration was 4 min.Electro-spray ionization ion source and multiple reaction monitoring mode were used to detect the positive ions.The established method was used to determine the plasma concentration of voriconazole and voriconazole N-oxide at every time point after intragastric administration of 33 mg·kg^(-1) voriconazole to rats and analyze its pharmacokinetic process in rats.Results:The established method showed good linearity over the range of 0.5-500 ng·ml^(-1)(r>0.9990).The lower limits of quantification both were 0.5 ng·ml^(-1).The pharmacokinetic parameters of voriconazole and voriconazole N-oxide were determined by the established method.The AUC_(0-t) values were(54.66±20.30)μg·ml^(-1)·h and(10.88±5.30)μg·ml^(-1)·h;AUC_(0-∞)were(54.66±20.30)μg·ml^(-1)·h and(13.50±2.10)μg·ml^(-1)·h;CL were(4.19±0.90)L·min^(-1)·kg^(-1) and(18.98±3.70)L·min^(-1)·kg^(-1);C_(max) were(19.14±11.90)μg·mL^(-1) and(0.95±0.20)μg·mL^(-1);t_(1/2) were(5.89±6.10)h and(6.11±3.20)h.Conclusion:The method is rapid,simple,accurate and reproducible,which is suitable for the pharmacokinetic study of voriconazole and voriconazole N-oxide in rats.