超高效液相色谱-四极杆线性离子阱串联质谱法测定他达拉非原料药中潜在基因毒性杂质的含量

Determination of genotoxic impurity in tadalafil raw materials by UPLC-q/LIT-MS

目的建立超高效液相色谱-四极杆线性离子阱串联质谱法测定他达拉非原料药中潜在的基因毒性合成工艺杂质T2的含量。方法采用Waters ACQUITY UPLC-AB SCIEX QTRAP 5500液质联用仪,ACQUITY UPLC HSS T3(2.1 mm×100 mm,1.8μm)色谱柱,以0.02 mol·L^(-1)乙酸铵(含0.1%乙酸)-乙腈为流动相进行梯度洗脱,流速0.35 mL·min^(-1),进样量5μL,柱温25℃。ESI正离子多反应监测(MRM)模式测定,离子对为m/z 427.1→135.0,外标法定量。结果该杂质在0.10~80 ng·mL^(-1)内与峰面积线性关系良好。检测限和定量限分别为25 pg·mL^(-1)和80 pg·mL^(-1),加样回收率在90%~110%,精密度、稳定性试验的RSD值均小于10%,说明该方法适用于他达拉非潜在基因毒性杂质T2的检测。含量测定结果显示,TDLF01、TDLF02原料药中T2杂质含量低于检测限,符合毒理学关注阈值的要求。结论该方法快速、灵敏、准确,可用于他达拉非原料药中潜在基因毒性杂质T2的含量测定。

Objective To establish an UPLC-q/LIT-MS method for the determination of intermediate impurity T2 produced in the synthesis of tadalafil raw materials.Methods The separation was operated on an ACQUITY UPLC HSS T3 column(2.1 mm×100 mm,1.8μm)with 0.02 mol·L^(-1)ammonium acetate including 0.1%acetic acid(A)and acetonitrile(B)as the mobile phase at Waters ACQUITY UPLC-AB SCIEX QTRAP 5500 platform.The flow rate was 0.35 mL·min^(-1).The injection volume was 5μL.The column temperature was 25℃and the data were acquired in ESI positive ion mode.The scan mode was MRM and the quantitative transition was m/z 427.1→135.0.Results The linear concentration range was from 0.10 to 80 ng·mL^(-1),the limit of detection was 25 pg·mL^(-1)and the limit of qualification was 80 pg·mL^(-1).The average recovery was at 90%~110%.The RSD of precision and stability was less than 10%.The content of T2 impurities in TDLF01 and TDLF02 was below the limit of detection,which met the TTC threshold.Conclusion The method is fast,sensitive and accurate,which can reliably control the quality of the intermediate impurity in tadalafil raw materials.