LC-MS/MS法体外测定肝匀浆液中乌头碱及其代谢产物

目的: 建立测定大鼠肝匀浆液中乌头碱含量的LC-MS/MS方法,评价附子甘草配伍前后在大鼠肝组织匀浆液中的体外温孵代谢情况。 方法: 对LC-MS/MS法进行验证,并测定不同温孵时间点甘草附子合煎液、附子单煎液加空白大鼠肝匀浆中乌头碱的含量及附子单煎液加甘草诱导后大鼠肝匀浆中乌头碱的含量。 结果: 建立的LC-MS/MS方法线性关系、精密度、准确度和灵敏度都符合样品分析测试要求。体外温孵6 h后,附子单煎液加空白组代谢率为10.97%,附子甘草合煎液加空白组代谢率为25.42%,附子单煎液加甘草诱导组代谢率为18.99%;体外温孵22 h后,附子单煎液加空白组代谢率为13.05%,附子甘草合煎液加空白组代谢率为47.01%,附子单煎液加甘草诱导组代谢率为32.13%。 结论: 所建立的LC-MS/MS方法用于大鼠肝匀浆中乌头碱的含量测定,简单、准确、灵敏。甘草与附子配伍后能加快乌头碱在肝脏中的代谢速度,且甘草与附子同时给药比甘草诱导后再给附子药液其乌头碱的代谢速度更快。

Objective: A high performance liquid chromatography-triple-quadrupole tandem mass spectrometry (LC-TQ-MS/MS) method was established and validated for the analysis of aconitine in rat liver homogenate, by which the effect of the compatibility of Radix Aconiti Laterlis (RAL) and Radix Glycyrrhizae (RG) on the liver metabolism of aconitine in vitro was investigated. Method: Three groups were designed: blank liver homogenate incubated with RAL extract (GroupⅠ), blank liver homogenate incubated with RAL and RG extract (GroupⅡ), and RG extract induced liver homogenate incubated with RAL extract (Group Ⅲ). At the different incubating time, the contents of aconitine in three groups of incubation liquids were determined by LC-MS/MS. Results: After incubating for 6 h, the metabolic rates of aconitine in groupⅠ, Ⅱ, Ⅲ were 10.97%, 24.42% and 18.99%, respectively. After incubating for 22 h, the data were 13.05%, 47.01% and 32.13%, respectively. Conclusion: The established LC-MS/MS method was sensitive and accurate for the quantification of aconitine in liver homogenate. When RAL was combined with RG, the liver metabolic rate of aconitine in vitro was accelerated. And if RAL and RG were applied simultaneously, the metabolic rate was higher than that when applied individually.