LC-MS/MS法测定Beagle犬血浆中美金刚的质量浓度

Determination of memantine hydrochloride tablets in Beagle dog by LC-MS/MS

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 develop an LC-MS/MS method to determine memantineand applied it to pharmacokinetics of memantine hydrochloride tablet in Beagle dogs. Methods Protein precipitation technique was utilized to treat plasma samples, amantadine was selected as internal standard, chromatographic separation was operated by usingGrace Altima HP C18 （50 mm × 2. 1 mm,5 μm）column, and water：formic acid（ V： V = 100： 0. 1 ）-acetonitrile： formic acid （ V： V = 100： 0. 1 ） was selected as mobile phase at gradient eluted system, the flow rate was 0. 35 mL· min^-1, quantification was performed using multiple reaction monitoring （MRM） of memantine using electrosprayionization source and positive ion switching mode. MRM transitions of m/z 179.8→m/z 162. 8 ,m/z 152.6→m/z 135.7 were selected for memantineand amantadine （IS） for quantification, respectively. 4 healthy male Beagle dogs were enrolled following a singleoral administration 25 mg of memantine hydrochloride tablets, the main pharmacokinetic parameters of memantine were calculated by WinNonlin 6. 1 software. Results The linearrange was 0. 001 -0.2 mg· L^- 1 for memantine, the lower limit of quantitation was 1 μg · L^-1. The intra-day and inter-day precision was between 2. 1% and 10. 8 %, and the accuracy （relative error）was between -3.5 % and 2.9%, the recovery was between 94.9% and 96. 8%, the main pharmacokinetics parameters following a single oral administration 25 mg of memantine hydrochloride tablet in Beagle dogs were as follows, t1/2 was （ 11.0 ±4.7）h,tmax was （9.0 ±2.0） h,ρmax was （0. 078 ±0. 024）mg·L^-1 ,AUC0-t was （1. 350 ±0. 295） mg·h·L^-1 , AUC0-∞was （ 1. 396 ±0. 306 ） mg· h· L ^- 1. Conclusions This validated method is within range of analytical method guidelines and can be applied to pharmacokinetics of memantine.