LC/MS/MS测定大鼠尿液中吴茱萸碱和吴茱萸次碱的方法学研究

Determination of Evodiamine and Rutaecarpine in Rat Urine by LC/MS/MS

目的:建立一种快速测定大鼠尿液中吴茱萸碱和吴茱萸次碱的定量方法。方法:用三重四级杆串联质谱(MS/MS)作为HPLC的检测器,其中MS/MS使用了多反应检测(MRM)扫描方式。分别选择304.2→134.2和288.2→244.2离子对m/z作为MRM检测的离子对;流动相:甲醇-水(85∶15),内含5mol/L甲酸铵;色谱柱:Agilent Zorbax XDB-C18,以测定大鼠尿液中吴茱萸碱和吴茱萸次碱为例,对此方法进行了应用。结果:方法的线性范围为吴茱萸碱=5.615-1437.5ng/ml(r=0.999),检测限1.87ng/ml,吴茱萸次碱=10.26-1314.25ng/ml(r=0.998),检测限5.13ng/ml。结论:此法灵敏、准确,可用于含茱萸碱和吴茱萸次碱的药物代谢研究。

Objective：To establish a method for quantitative analysis of evodiamine and rutaecarpine in rat urine rapidly.Methods：A triple-quadrupole tandem mass spectrometry was used as the detector of HPLC.As to MS/MS,multi-reactions monitoring（MRM）scan mode was employed.Among the product ions of evodiamine and rutaecarpine,m/z-134.2 and 244.2 are the most abundant in intensity,thus the parent-daughter ion pairs of m/z 304.2→134.2 and 288.2→244.2 were selected as MRM ions pairs.MS/MS conditions were optimized to achieve highest sensitivity.The mobile phase of HPLC was methanol-water（85：15）containing 5mM ammonium formate.The analytical column was Agilent Zorbax XDB-C18.The flow rate of HPLC was 0.5ml/min.The areas of ion flow peaks were used to determine the content of evodiamine and rutaecarpine in the rat urine.Results：The standard curves for evodiamine and rutaecarpine showed good linearity over ranges of 5.615-1437.5ng/ml（r=0.999）and 10.26-1314.25ng/ml（r=0.998）,and the detection limit for evodiamine and rutaecarpine were 1.87ng/ml and 5.13ng/ml respectively.Conclusion：This method is highly sensitive,fast and very accurate.Therefore,it is possible to be applied to study the metabolism of evodiamine and rutaecarpine.