目的 建立液相色谱-串联质谱法(LC-MS/MS)检测人血浆中阿兹夫定(azvudine, AZV)的浓度,并应用于新型冠状病毒感染患者的临床样本测定。方法 血浆经Ostro^TM板除蛋白及磷脂后,以¹⁵N₄-次黄嘌呤为内标,采用LC-MS/MS测定。色谱柱采用HILIC Silica柱,以乙腈(0.1%甲酸)-10 mmol·L^-1甲酸铵(0.1%甲酸及5%乙腈)为流动相进行梯度洗脱,流速0.30 mL·min^-1,柱温40 ℃,进样量0.50 μL。采用电喷雾离子源,以多反应监测模式进行正离子扫描,用于定量分析的离子对为m/z 287.3→112.0(AZV)、m/z 141.0→113.0(内标)。结果 血浆中AZV在0.10~20.00 ng·mL^-1内线性良好,定量下限日内、日间相对标准偏差(RSD)均不大于4.46%,相对误差为12.00%~15.13%;低、中、高质控样本日内、日间RSD均不高于2.88%,相对误差为3.10%~12.11%。平均提取回收率为75.69%,基质效应及残留均不影响待测物的准确定量。该方法经验证后,成功应用于慢性透析合并新型冠状病毒感染患者血浆中AZV浓度的测定。结论 本试验所建立的LC-MS/MS简单、灵敏、准确,可用于AZV的治疗药物监测。

OBJECTIVE To establish an LC-MS/MS method for the determination of azvudine (AZV) concentration in plasma samples from patients with corona virus disease 2019 (COVID-19). METHODS After protein precipitation and phospholipid removal with 96-well Ostro^TM plate, AZV in human plasma was determined by LC-MS/MS method. ¹⁵N₄-hypoxanthine was used as the internal standard (the IS). Gradient elution was performed on a HILIC silica column. The gradient mobile phase consisted of acetonitrile (0.1% formic acid) and 10 mmol·L^-1 ammonium formate (0.1% formic acid and 5% acetonitrile) at a flow rate of 0.3 mL·min^-1. The column temperature was maintained at 40 ℃ and the injection volume was 0.50 μL. An ESI source was used on positive mode. MRM transitions were m/z 287.3→112.0 (AZV) and m/z 141.0→113.0 (the IS). RESULTS The assay was linear within the range of 0.10-20.00 ng·mL^-1. Intra-and inter-day precision for LLOQ was less than 4.46% and the relative error was 12.00%-15.13%. For quality control samples, the intra-and inter-day precisions were less than 2.88% and the relative error was 3.10%-12.11%. The average extraction recovery was 75.69%. The matrix effect and carryover were negligible. After validation, the method was applied to the determination of AZV in plasma of chronic dialysis patients with COVID-19. CONCLUSIONS The established LC-MS/MS method is simple, sensitive and accurate, and it can be applied to the therapeutic drug monitoring of AZV in clinical practice.