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目的建立LC-MS/MS法测定Beagle犬血浆中美金刚的质量浓度,并将此方法应用于盐酸美金刚片在Beagle犬体内的药动学研究。方法采用蛋白沉淀法处理血浆样品,以金刚烷胺为内标物质(IS),采用Grace Altima HP C_(18)(50 mm×2.1 mm,5μm)色谱柱,流动相为体积分数0.1%甲酸水溶液-体积分数0.1%甲酸乙腈溶液,梯度洗脱,流速为0.35 m L·min~(-1)。离子源为电喷雾电离(ESI)源,采用正离子化方式测定,采用多反应监测(multiple reaction monitoring,MRM)模式检测,用于定量分析的离子反应为m/z 179.8→m/z 162.8(美金刚),m/z 152.6→m/z 135.7(金刚烷胺,内标物质)。4只健康雄性Beagle犬单次经口给予盐酸美金刚片25 mg,用Win Nonlin 6.1软件以非房室模型计算药动学参数。结果血浆中美金刚线性范围为0.001~0.2 mg·L~(-1),定量下限为1μg·L~(-1),日内和日间精密度在2.1%~10.8%内,准确度在-3.5%~2.9%内,提取回收率在94.9%~96.8%内。Beagle犬单次经口给予盐酸美金刚片后,主要的药动学参数t_(1/2)为(11.0±4.7)h,t_(max)为(9.0±2.0)h,ρ_(max)为(0.078±0.024)mg·L~(-1),AUC_(0-t)为(1.350±0.295)mg·h·L~(-1),AUC_(0-∞)为(1.396±0.306)mg·h·L~(-1)。结论所建立的定量分析方法符合生物样品分析方法指导原则要求,适用于盐酸美金刚片的药动学研究。
Objective To establish a method for the determination of memantine in Beagle dogs by LC-MS / MS. The method was applied to the pharmacokinetics of memantine hydrochloride in Beagle dogs. Methods Plasma samples were processed by protein precipitation method. Amantadine was used as internal standard substance (IS), and the column was on a Grace Altima HP C 18 (50 mm × 2.1 mm, 5 μm) column. The mobile phase consisted of 0.1% - volume fraction of 0.1% formic acid in acetonitrile, gradient elution, flow rate of 0.35 m L · min -1. The ion source was electrospray ionization (ESI) source and was detected by positive ionization mode and multiple reaction monitoring (MRM) mode. The ion reaction for quantitative analysis was m / z 179.8 → m / z 162.8 ( Memantine), m / z 152.6 → m / z 135.7 (amantadine, internal standard substance). Four healthy male Beagle dogs were orally given 25 mg memantine hydrochloride tablets and their pharmacokinetic parameters were calculated in a non-compartmental model using Win Nonlin 6.1 software. Results The linear range of memantine in plasma was 0.001-0.2 mg · L -1, the lower limit of quantitation was 1 μg · L -1, the intra- and inter-day precision was within 2.1% -10.8%, and the accuracy was between - 3.5% ~ 2.9% within the recovery rate of 94.9% ~ 96.8%. The main pharmacokinetic parameters of Beagle dogs were (11.0 ± 4.7) h, t max (9.0 ± 2.0) h and ρ max (0.78 ± 0.024) mg · L -1, AUC 0-t was 1.350 ± 0.295 mg · h · L -1, and AUC 0-∞ was 1.396 ± 0.306 mg / · H · L ~ (-1). Conclusion The established quantitative analysis method conforms to the guidelines of biological sample analysis method and is suitable for pharmacokinetic study of memantine hydrochloride tablets.