高效液相色谱串联电喷雾质谱法测定大鼠血浆中山莨菪碱

Determination of anisodamine in plasma of rat by HPLC-MS

目的建立大鼠血浆中山莨菪碱的测定方法,并研究山莨菪碱的大鼠药动学。方法采用液相色谱串联电喷雾离子阱质谱法测定大鼠血浆中山莨菪碱。大鼠血浆样品经甲醇2次沉淀蛋白并萃取后,以甲醇-2mmol/L醋酸铵缓冲液(用甲酸调pH3.5)(70:30)为流动相,用AgilentZorbaxEclipseXDB-C_(18)色谱柱分离,通过电喷雾离子阱质谱,以选择离子反应监测方式进行检测。用于定量分析的离子反应分别为m/z306→140(山莨菪碱)和m/z290→124(内标,阿托品)。结果山莨菪碱线性范围为10-5000μg/L,r=0.9990,最低检测限为2μg/L,日内、日间测定值RSD小于4.2%。结论该法快速灵敏,准确度高,适用于大鼠体内山莨菪碱的药动学研究。

Objective To establish a sensitive and specific method for the determination of anisodamine in plasma of rat and to study pharmacokinetics of anisodamine in plasma of rat. Methods HPLC- MS was used to determine the concentration of anisodamine in plasma of rat with atropine as the internal standard. The atropine was isolated from plasma by protein precipitation and extraction with methanol twice, then chromatographed by using an Agilent Zorbax Eclipse XDB-C18 column. The mobile phase consisted of methanol-2 mmol/L ammonium acetate solution （adjusted pH value to 3.5 with formic acid） 70 ： 30. Electrospray ionization （ESI） source was applied and operated in the positive ion mode. Selected reaction monitoring （SRM） mode with the transitions of m/z 306→140 and m/z 290→124 was used to quantify anisodamine and the internal standard, respectively. Results The linear range for anisodamine was 10- 5 000μg/L （r = 0. 999 0）, the limit of detection for anisodamine was 2μg/L, inter- and intra-day RSD was less than 4.2%. Conclusion The method offers the advantages of high sensitivity and accuracy for the determination of anisodamine compared with the previously reported methods and is suitable for the pharmacokinetic study on anisodamine in rat in vivo.