UPLC-MS/MS法测定大鼠血浆中抗耐多药结核活性化合物macozinone(PBTZ 169)浓度及其药代动力学研究

Determination of the active compound against multi-resistant tuberculosis macozinone (PBTZ 169) concentration in rat plasma by UPLC-MS/MS method and its pharmacokinetics

目的:建立UPLC-MS/MS法测定抗耐多药结核新型活性化合物macozinone (PBTZ 169)在大鼠血浆中的浓度,并应用于大鼠体内的药代动力学研究。方法:以卡马西平为内标,经乙腈沉淀蛋白后,采用XBridge?BEH C_(18)(100 mm×2.1 mm, 2.5μm)色谱柱,以乙腈-0.1%甲酸水为流动相,进行梯度洗脱分离,流速0.5 mL·min^(-1),进样量10μL;采用电喷雾离子源(ESI),正离子扫描,分别以m/z 457.2→344.1和m/z 237.2→194.2作为PBTZ 169和内标(卡马西平)的质谱检测条件。大鼠灌服20 mg·kg^(-1)PBTZ 169后,不同时间点取样测定其血浆浓度,计算药代动力学参数并进行统计分析。结果:PBTZ 169质量浓度在0.7~740.0 ng·mL^(-1)范围内线性关系良好(r>0.996 0);PBTZ 169和内标的提取回收率在99.5%~110.2%;日内、日间的RSD范围为1.3%~4.2%;PBTZ 169血浆样品在室温下放置6 h,血样处理后自动进样器放置24 h,-70℃贮存15 d,-70℃反复冻融3次的稳定性试验的RSD均<15%。大鼠口服灌胃PBTZ 169后,PBTZ 169在大鼠体内主要药代动力学参数:Cmax为(374.8±116.3) ng·mL^(-1),Tmax为(0.8±0.1) h, AUC0~∞为(1 180.0±383.5) ng·h·mL^(-1),T1/2为(3.7±0.7) h, MRT0~∞为(4.7±1.2) h。结论:该方法简便、快捷、灵敏,适用于PBTZ 169的药代动力学研究。

Objective:To establish a rapid and accurate method of UPLC-MS/MS for the determination of macozinone(PBTZ 169) concentration, a novel active compound against multi-resistant tuberculosis, in rat plasma and to study its pharmacokinetics in rats. Methods: Carbamazepine was used as internal standard. After the protein wasprecipitated by acetonitrile, the plasma samples were separated on the XBridge? BEH C_(18)(100 mm×2.1 mm, 2.5 μm) chromatographic column through gradient elution by acetonitrile and 0.1% formic acid as mobile phase. The flow rate was 0.5 mL·min^(-1)and sample volume was 10 μL. Then the analytes were detected by the electrospray ionization(ESI) sourcein with positive ion scanning. The detection conditions of PBTZ 169 and internal standard(carbamazepine) by mass spectrometry were m/z 457.2→344.1 and m/z 237.2→194.2, respectively. After the rats were given 20 mg·kg^(-1)PBTZ 169 and the plasma concentration of PBTZ 169 was detected at different time points. The pharmacokinetic parameters were calculated and statistically analyzed. Results: The mass concentration of PBTZ 169 had a good linear relationship between 0.7 and 740.0 ng·mL^(-1)(r=0.996 0). The extraction recovery of PBTZ 169 and internal standard were between 99.5% and 110.2% and the RSDs of intra day and inter day were ranged from 1.3% to 4.2%. PBTZ 169 plasma samples were placed at room temperature for 6 h, the automatic sampler was placed for 24 h after the blood sample was processed, refrigerated at-70 ℃ for 15 d and repeatedly freeze-thaw at-70 ℃ for 3 times. Finally, the RSDs were all lower than 15%. After oral intragastric administration of PBTZ 169 to rats, the main pharmacokinetic parameters of PBTZ 169 in rats were as follows: Cmaxwas(374.8±116.3) ng·mL^(-1), Tmaxwas(0.8±0.1) h, AUC0~∞was(1 180.0±383.5) ng·h·mL^(-1), T1/2was(3.7±0.7) h, MRT0~∞was(4.7±1.2) h. Conclusion: This method is simple, rapid, sensitive and suitable for pharmacokinetic study of PBTZ 169.