目的: 建立超高液相色谱-大气压化学电离源三重四极杆质谱法(UPLC-APCI MS/MS)检测克拉霉素中3个N-亚硝胺类化合物[N-亚硝基乙基异丙基胺(NIPEA)、N-亚硝基二异丙胺(NDIPA)和N-亚硝基二丁胺(NDBA)。方法: 采用ACQUITY UPLC HSS T3色谱柱(100 mm×2.1 mm,1.8 μm),以甲醇-0.1%甲酸水溶液为流动相,梯度洗脱,流速0.4 mL·min-1;采用大气压化学电离(APCI),多反应监测(multiple reaction monitoring,MRM),正离子扫描模式。结果: 测得NIPEA、NDIPA质量浓度在18~120 μg·L-1内呈现良好的线性关系(r=0.999),NDBA质量浓度在7.8~52 μg·L-1内呈现良好的线性关系(r=0.999 1),重复性试验的RSD为2%~3%,方法回收率为88%~109%,NIPEA、NDIPA方法定量限(S/N不低于10)为9.0 μg·L-1,NDBA方法定量限(S/N不低于10)为3.9 μg·L-1,溶液在室温放置24 h内稳定。对2家国内生产企业各3批克拉霉素样品进行检测,NIPEA、NDIPA、NDBA均低于检测限。结论: 本试验建立了克拉霉素中N-亚硝胺类化合物NIPEA、NDIPA、NDBA的检测方法,具有灵敏度高,测定结果准确,专属性强等诸多优点,可应用于临床治疗药物的监测。
Objective: To establish an UPLC-APCI MS/MS method for the determination of three N-nitrosamines (NIPEA、NDIPA、NDBA) in clarithromycin. Methods: An ACQUITY UPLC HSS T3(100 mm×2.1 mm, 1.8 μm) was adopted, the mobile phase consisted of methanol-0.1% formic acid solution with gradient elution, at a flow rate of 0.4 mL·min-1; atmospheric-pressure chemical ionization (APCI), multiple reaction monitoring (MRM), positive ion scanning mode were used. Results: The measured mass concentrations of NIPEA and NDIPA had a good linearity in the range of 18-120 μg·L-1(r=0.999), and the mass concentration of NDBA had a good linearity in the range of 7.8-52 μg·L-1(r=0.999 1). The RSD of all repeatability tests were 2%-3%, the recovery rates were 88%-109%. The LOQ(S/N was about 10) of the methods for NIPEA and NDIPA were 9.0 μg·L-1 and that for NDBA was 3.9 μg·L-1. The solution was stable at room temperature in 24 h. Three batches of clarithromycin samples from two domestic manufacturers were tested, and NIPEA, NDIPA and NDBA were below LOD. Conclusion: This study established a method for the determination of N-nitrosamines NIPEA, NDIPA and NDBA in clarithromycin with the advantages of high sensitivity, accurate determination results, strong specificity and so on. It can be applied to the monitoring of clinical therapeutic drugs.
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