目的:建立超高效液相色谱-串联质谱(UPLC-MS/MS)方法测定大鼠血浆中刺五加提取液中7-羟基香豆素、秦皮素、异嗪皮啶、东莨菪内酯4个成分的含量,评价大鼠体内的药代动力学行为和生物利用度。方法:采用Thermo Scientific Hypersil GOLD aQ 色谱柱(100 mm×2.1 mm, 1.9 μm),以0.1%甲酸-水(A)和0.1%甲酸-乙腈(B)(82∶18)为流动相,流速0.30 mL·min-1,柱温40 ℃;质谱正离子模式扫描,进样量5 μL;选取健康SD大鼠,单次灌胃10 mL·kg-1(相当于原药1 g·kg-1剂量)的刺五加提取液,测定给药后不同时间间隔的待测物质血浆浓度,利用DAS软件,经非房室模型拟合计算药代动力学参数。结果:7-羟基香豆素(r=0.999 4)、秦皮素(r=0.998 9)、异嗪皮啶(r=0.999 3)、东莨菪内酯(r=0.998 4)质量浓度在0.05~55 μg·mL-1范围内线性关系良好,精密度RSD均<15%,回收率均在85%~115%。绝对生物利用度均在59%~78%,相对生物利用度均在80%~87%。提取回收率、基质效应和稳定性均满足相关要求。结论:大鼠单次灌胃刺五加提取液后,4个待测物质均被大鼠吸收和消除,方法验证结果符合生物样本分析方法指导原则,可用于评价刺五加提取液在大鼠体内的药代动力学行为和生物利用度。
Objective: To establish a ultra-high performance liquid chromatography-mass spectrum in series method for the quantification of 7-hydroxycoumarin, fraxetin, isofraxidin and scopoletin in the aqueous extract of Acanthopanacis Senticosi Radix et Rhizoma Seu Caulis in rat plasma, and to evaluate the pharmacokinetic behavior and bioavailability in rats. Methods: The chromatography was performed on Thermo Scientific Hypersil GOLD aQ (100 mm×2.1 mm, 1.9 μm)column with 0.1% aqueous formic acid -0.1% acetonitrile formic acid solution (82∶18) as the mobile phase at the flow rate of 0.30 mL·min-1, and column temperature of 40 ℃. Mass spectrometry positive ion mode scan was used, and sample size was 5 μL. Healthy SD rats were selected for a single gavage of 10 mL·kg-1 of Acanthopanacis Senticosi Radix et Rhizoma Seu Caulis water extract (equivalent to the dose of 1 g·kg-1 of the original drug). Plasma concentration of the substances was determined at different time intervals after administration, and the pharmacokinetic parameters were calculated using DAS software by non-atrioventricular model fitting. Results: The methodological results showed that 7-hydroxycoumarin (r=0.999 4), fraxetin (r=0.998 9), isofraxidin (r=0.999 3) and scopoletin (r=0.998 4) had good linearity in the range of 0.05-55 μg·mL-1, and RSDs of precision of the four substances were all less than 15%. The recovery was 85%-115%. The absolute bioavailability ranged from 59% to 78%, and the relative bioavailability ranged from 80% to 87%. The extraction recovery, matrix effect and stability met the relevant requirements. Conclusions: After a single gavage of the aqueous extract of Acanthopanacis Senticosi Radix et Rhizoma Seu Caulis in rats, the four substances to be tested are absorbed and eliminated by rats. The method validation results are in line with the guiding principles of biological sample analysis methods, and can be applied to evaluate the pharmacokinetic behavior and bioavailability of the aqueous extract of Acanthopanacis Senticosi Radix et Rhizoma Seu Caulis in rats.
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