目的: 建立一种采用LC-QTOF/MS同时测定他达拉非原料中3个潜在基因毒性杂质:氯乙酰氯(JD-1)、(1R,3R)-1-(1,3-苯并二氧戊环-5-基)-2-(氯乙酰基)-2,3,4,9-四氢-1H-吡啶并[3,4-b]吲哚-3-羧酸甲酯(JD-2)和(1S,3R)-1-(1,3-苯并二氧戊环-5-基)-2-(氯乙酰基)-2,3,4,9-四氢-1H-吡啶并[3,4-b]吲哚-3-羧酸甲酯(JD-3)含量的方法。方法: 采用苯胺为衍生化试剂,对样品进行前处理,采用YMC-PackODS-AQ色谱柱进行分离,采用乙腈和水体系为流动相,进行梯度洗脱,流速为1.0 mL·min-1。质谱采用电喷雾离子源,负离子模式下选择[M-H]-m/z 168.050 0和[M-H]-m/z 425.085 5作为JD-1和JD-2、JD-3的定量离子。结果: 3个潜在基因毒性杂质在37.5～112.5 ng·mL-1范围内线性关系良好(r＞0.990 0),JD-1、JD-2和JD-3的定量限(LOQ)分别为3.8、7.3和4.1 ng·mL-1,3个潜在基因毒性杂质的平均回收率均在80%~120%范围内,重复性试验含量RSD均小于10%。3批原料中JD-2的含量分别为19.5、22.1、20.0 μg·g-1,JD-3的含量分别为21.9、10.1、16.3 μg·g-1,3批原料中均未检出JD-1。结论: 该方法结果可靠,可用于他达拉非原料中3个潜在基因毒性杂质JD-1、JD-2、JD-3的含量测定。
Objective: To establish a method was established for the determination of three potential genotoxic impurities, chloroacetyl chloride(JD-1) and (1R, 3R)-1-(1, 3-benzodioxol-5-yl)-2-(chloroacetyl)-2, 3, 4, 9-tetrahydro-1H-pyrido[3, 4-b] indole-3-carboxylic acid methyl ester(JD-2) and (1S, 3R)-1-(1, 3-benzodioxol-5-yl)-2-(chloroacetyl)-2, 3, 4, 9-tetrahydro-1H-pyrido[3, 4-b]indole-3-carboxylic acid methyl ester(JD-3) in tadalafil by LC-QTOF/MS. Methods: The samples were pretreated with aniline as a derivatization reagent. The separation was performed on a YMC-Pack ODS-AQ column with the mobile phase consisting of water and acetonitrile by gradient elution at a flow rate of 1.0 mL·min-1. Electronic spray ion (ESI) in negative ionization mode was performed. The [M-H]- ion(m/z 168.050 0) was selected as thequantitative ion of JD-1 and the [M-H]- ion(m/z 425.085 5) was selected as the quantitative ion of JD-2 and JD-3. Results: The calibration curves of three potential genotoxic impurities were in good linearity (r>0.990 0) within the range of 37.5-112.5 ng·mL-1. The lower limits of quantification (LOQ) of JD-1, JD-2 and JD-3 were 3.8, 9.9 and 4.1 ng·mL-1, respectively. The average recoveries of the three impurities were in the range of 80%-120% and the RSDs of repeatability tests were less than 10.0%. Conclusion: The method is accurate and reliable to determine three potential genotoxic impurities JD-1, JD-2 and JD-3 in tadalafil.
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