目的 建立超高效液相色谱串联质谱法测定大鼠血浆中薯蓣皂苷元浓度。方法 以丹参酮ⅡA为内标。血浆经甲醇沉淀蛋白后,采用色谱柱为Phenomenex kinetex xb C18(2.1 mm×50 mm,2.6 μm),以甲醇(含0.1%甲酸)(A)-0.1%甲酸(B)为流动相,梯度洗脱,流速为0.2 mL·min-1,柱温为40 ℃,进样量为5 μL;以电喷雾(ESI)离子源,在正离子电离模式下,采用多反应监测(MRM)的扫描模式进行测定。薯蓣皂苷元和丹参酮ⅡA的MRM扫描离子对分别为m/z 415.2→271.1和m/z 295.1→249.1。结果 血浆中薯蓣皂苷元质量浓度在10~500 ng·mL-1内线性关系良好(r=0.998 3);最低定量限为10 ng·mL-1;准确度为96.1%~102.3%;日内和日间精密度均小于15%;平均提取回收率为73.8%~75.2%;基质效应为85.8%~91.7%。内标的平均提取回收率为83.8%,基质效应为92.4%。大鼠灌胃给予薯蓣皂苷元(100 mg·kg-1)后,薯蓣皂苷元在大鼠体内的达峰浓度为(344.067±34.48)ng·mL-1,达峰时间为(4.167±2.041)h,半衰期为(14.85±10.53)h,血药浓度-时间曲线下面积为(4 965.648±1 036.129)μg·h·L-1。结论 该方法专属性强,样品处理方便,灵敏度高,检测时间短,适用于薯蓣皂苷元在大鼠体内的药动学研究。
Abstract
OBJECTIVE To develop and validate a sensitive and specific ultra-performance liquid chromatography-tandemmass spectrometric (LC-MS/MS) method for the assay of diosgenin in rat plasma. METHODS Tanshinone ⅡA was employed as internal standard. Diosgenin was determined after the methanol-mediated plasma protein precipitation. The separation was performed on the Phenomenex kinetex xb C18 column (2.1 mm×50 mm,2.6 μm) gradiently eluted with the mobile phase consisting of methanol(containing 0.1% formic acid)-0.1% aqueous formic acid. The flow rate was 0.2 mL·min-1, the column temperature was maintained at 40 ℃, and the injection volume was 5 μL. A triple quadrupole mass spectrometer equipped with electrospray ionization source was used as detector in a positive ion mode. Multiple reaction monitoring (MRM) mode was applied with the transition of m/z 415.2→271.1 and m/z 295.1→249.1 for diosgenin and internal standard, respectively. RESULTS For diosgenin the standard curve was linear from 10 to 500 ng·mL-1(r=0.998 3), the limit of quantitative limit was 10 ng·mL-1, the intra- and inter-assay variabilities were below 15%, the accuracies were between 96.1% and 102.3%, the average extract recoveries ranged from 73.8% to 75.2%, and the matrix effects was between 85.8% and 91.7%. For the internal standard, the extract recovery and matrix effects were 83.8% and 92.4%, respectively. The rats were administered orally with diosgenin (100 mg·kg-1). The peak concentration of diosgenin was (344.067±34.48) ng·mL-1, the time for peak concentration was (4.167±2.041) h, the half-time was (14.85±10.53) h, and the area under concentration-time curve from zero to 72 h was (4 965.648±1 036.129) μg·h·L-1. CONCLUSION This assay is specific, simple, sensitive and rapid, which can be applied in the pharmacokinetic study of diosgenin in rats.
关键词
薯蓣皂苷元 /
血药浓度 /
药动学 /
超高效液相色谱串联质谱法
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Key words
diosgenin /
plasma concentration /
pharmacokinetics /
UPLC-MS/MS
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中图分类号：
R917
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参考文献
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脚注
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基金
河北省自然科学基金资助项目(H2014406036);河北省高等学校科学技术研究资助项目(QN2015127);河北省高校重点学科建设资助项目(冀教高[2013]4号)
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