UPLC-MS/MS法同时检测大鼠血浆中参芪扶正注射液主要成分的含量及其药动学

Content determination and pharmacokinetic of main ingredients of Shenqi Fuzhen injection in rat plasma

目的:建立超高效液相-三重四极杆质谱联用(UPLC-MS/MS)法同时测定大鼠血浆中黄芪甲苷、毛蕊异黄酮-7-O-β-D葡萄糖苷和党参炔苷的浓度,研究参芪扶正注射液在大鼠体内的药动学。方法:血浆样品加入适量内标,50%甲醇-乙腈沉淀蛋白,离心后取上清进样。采用ACE Excel 1.7C_(18)柱(50mm×2.1mm,2.1μm),柱温:40℃,流速:0.6mL·min^(-1),流动相由0.1%甲酸水和0.1%甲酸-乙腈组成,采用梯度洗脱,分析时间2.5min。质谱采用ESI源,正离子检测。大鼠尾静脉注射参芪扶正注射液浓缩液,眼眶采血测定血浆中药物浓度,药动学参数以DAS 3.0软件处理。结果:黄芪甲苷、毛蕊异黄酮-7-O-β-D葡萄糖苷、党参炔苷的浓度分别在5~2 000ng·mL^(-1)、2~800ng·mL^(-1)、10~4 000ng·mL^(-1)内均呈良好线性关系(R≥0.991 5);日内、日间精密度良好,RSD均小于13.3%(n=6);提取回收率均在79.0%~97.1%之间,RSD均小于11.4%(n=6)。药动学结果表明,黄芪甲苷、党参炔苷和毛蕊异黄酮-7-O-β-D葡萄糖苷在大鼠体内的AUC0-t分别为(38.84±17.20)、(23.11±6.84)、(5.32±0.36)μg·min·L^(-1),t1/2分别为(56.44±28.25)、(47.48±9.54)、(10.01±4.42)min,CL分别为(0.15±0.06)、(1.18±0.47)、(0.47±0.11)mL·min-1·kg^(-1)。结论:本法简便、快速、灵敏度高、专属性好,可用于参芪扶正口服液大鼠血浆中黄芪甲苷、毛蕊异黄酮-7-O-β-D葡萄糖苷和党参炔苷的含量测定及药动学研究。

OBJECTIVE To establish an UPLC-MS/MS method for determination of astragaloside Ⅳ,calycosin-7-O-β-Dglucoside and lobetyolin and investigate their pharmacokinetics in rat plasma.METHODS The internal standard was added to the plasma samples,50% methanol-acetonitrile precipitation protein,centrifuged after the supernatant injection.All the separations were carried on an ACE Excel 1.7 C_（18） column（50 mm×2.1 mm,2.1μm）with the mobile phase of acetonitrile-water（both containing 0.1%formic acid）at gradient elution pumped at a flow rate of 0.6 mL·min-1 under 40 ℃ for 2.5 min.The analytes were detected on a triple-quadrupole mass spectrometer equipped with an ESI interface in a positive mode.RESULTSThe astragalosideⅣ,calycosin-7-O-β-D-glucoside and lobetyolin showed good linearity（R≥0.991 5）respectively within the ranges of 5-2 000 ng·mL-1,2-800 ng·mL-1,10-4 000 ng·mL-1.The intra-and inter-day RSDs were below 13.3%（n = 6）.The average recovery was within the range of 79.0%–97.1% with RSD below 11.4%（n=6）.The main pharmacokinetic parameters were as follows：the AUC0-tof astragaloside Ⅳ,lobetyolin,and calycosin-7-O-β-D-glucoside in rats were（38.84±17.20）,（23.11±6.84）,（5.32±0.36）μg·min·L-1 and t1/2 were（56.44±28.25）,（47.48±9.54）,（10.01±4.42）min,and CL were（0.15±0.06）,（1.18±0.47）,（0.47±0.11）mL·min-1·kg-1,respectively.CONCLUSION The method is rapid,sensitive and accurate for the quantification of astragaloside Ⅳ,calycosin-7-O-β-D-glucoside and lobetyolin and their pharmacokinetics in rat plasma,which provides a research basis for Shenqi Fuzheng injection in clinical practice.