超高效液相色谱-串联质谱法测定人血浆中盐酸氨溴索及其制剂的生物等效性评价

采用超高效液相色谱-串联质谱（UHPLC-MS/MS）技术，建立了快速、简单、灵敏的测定人血浆中盐酸氨溴索含量的方法，并用于盐酸氨溴索人体生物等效性预试验研究。取50 μL血浆样品，采用蛋白沉淀法处理，以盐酸氨溴索-d5为内标。采用Waters XBridge BEH C18色谱柱（50 mm×2.1 mm，2.5 μm），以0.1%（v/v）甲酸水-含0.1%（v/v）甲酸的甲醇为流动相，在0.4 mL/min流速下进行梯度洗脱。采用电喷雾电离（ESI）源以正离子模式进行MRM检测。结果显示，盐酸氨溴索在2~400 ng/mL范围内线性关系良好，相关系数（r）为0.998，准确度为97.1%~108.7%，精密度为1.0%~5.6%。将该方法用于6名健康受试者口服盐酸氨溴索受试制剂和参比制剂30 mg后血药浓度的测定，结果显示二者相对生物利用度为（102.3±14.8）%，血药浓度-时间曲线下面积（AUC_0-t、AUC_0-∞）和最大血药浓度（C_max）的90%置信区间均在80.0%~125.0%范围内，两种制剂生物等效。

Determination of ambroxol hydrochloride in human plasma by ultra high performance liquid chromatography-tandem mass spectrometry and bioequivalence evaluation of its preparation

A rapid, simple and sensitive ultra high performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) method was developed for the determination of ambroxol hydrochloride in human plasma, and bioequivalence of its preparation was evaluated. The 50 μL-plasma sample was treated with methanol for protein precipitation, while ambroxol-d5 was used as an internal standard (IS). The separation was carried out on a Waters XBridge BEH C18 column (50 mm×2.1 mm, 2.5 μm) by gradient elution at a flow rate of 0.4 mL/min, with 0.1% (v/v) formic acid aqueous solution and methanol containing 0.1% (v/v) formic acid as the mobile phases. The analyte was detected using an electrospray ionization source in positive ion multiple reaction monitoring (MRM) mode. The calibration curves were linear in the range of 2-400 ng/mL (r=0.998). The intra- and inter-run accuracies were 97.1%-108.7%, the intra- and inter-run precisions were 1.0%-5.6%. The method was applied to the determination of the plasma concentration of the six healthy subjects after the oral administration of 30 mg of test and reference preparations. The bioavailability was (102.3±14.8)%. The 90% confidence intervals of the test preparation's pharmacokinetic parameters were 80.0%-125.0% of the reference preparation's corresponding parameters. Thus, it is proved that the test preparation and reference preparation are bioequivalent.