AstraZeneca R&D, Mölndal, Sweden.
J Chromatogr B Analyt Technol Biomed Life Sci. 2010 Sep 1;878(25):2299-306. doi: 10.1016/j.jchromb.2010.06.018. Epub 2010 Jun 25.
Rapid and sensitive analytical methods using liquid chromatography with tandem mass spectrometry (LC/MS/MS) were developed for the determination of ticagrelor, the first reversible oral platelet P2Y(12) receptor inhibitor, and its metabolites AR-C124910XX and AR-C133913XX in human plasma. Ticagrelor and its metabolites were extracted using protein precipitation with acetonitrile. Chromatographic separations were performed on reversed phase columns and detection using atmospheric pressure chemical ionization (APCI). Ticagrelor and AR-C124910XX were analyzed in the same assay, with the internal standard, d7-ZD6140, on a C18 column using negative ionization; AR-C133913XX analyzed separately on a phenyl column using positive ionization. Full validation of the methods was performed including selectivity, lower limit of quantification, accuracy, precision stability and incurred sample reproducibility and incurred sample stability. Total analytical run time was short (2 min). Calibration curves were established in the range 5-5000ng/mL for ticagrelor, 2.5-2500 ng/mL for AR-C124910XX and 2-1000 ng/mL for AR-C133913XX. Lower limits of quantification for ticagrelor, AR-C124910XX and AR-C133913XX were determined to be 5, 2.5 and 2.0 ng/mL, respectively from 100 microL of human plasma. For ticagrelor, AR-C124910XX and AR-C133913XX, mean intra-batch accuracy was 91.9-109.0%, 86.8-109.2% and 100.5-112.0%, respectively; intra-batch precision was 4.0-8.4%, 5.2-16.9% and 3.9-12.3%, respectively. The methods were also applied to quantification of ticagrelor, AR-C124910XX and AR-C133913XX in rabbit, rat, mouse and marmoset, using 25 microL of animal plasma. A modified methodology was developed to quantify ticagrelor and AR-C124910XX in plasma from dog and cynomolgus monkey. Human incurred samples were found to generate consistent reproducibility and stability results. This method was successfully applied to determine plasma concentrations following administration of ticagrelor in human volunteers and patients, and animal safety evaluation studies. This validated methods has the advantages of being straightforward, robust and allows a fast throughput of samples.
建立了一种使用液相色谱-串联质谱（LC/MS/MS）快速灵敏测定替格瑞洛及其代谢物 AR-C124910XX 和 AR-C133913XX 的分析方法。替格瑞洛及其代谢物采用乙腈沉淀蛋白法提取。采用反相色谱柱分离，大气压化学电离（APCI）检测。替格瑞洛和 AR-C124910XX 在同一分析中进行分析，以内部标准 d7-ZD6140 在 C18 柱上采用负电离进行分析；AR-C133913XX 则在苯基柱上采用正电离进行分析。对方法进行了全面验证，包括选择性、定量下限、准确度、精密度、稳定性、重复性和稳定性。总分析时间短（2 分钟）。替格瑞洛、AR-C124910XX 和 AR-C133913XX 的校准曲线范围分别为 5-5000ng/mL、2.5-2500ng/mL 和 2-1000ng/mL。替格瑞洛、AR-C124910XX 和 AR-C133913XX 的定量下限分别为 5、2.5 和 2.0ng/mL，来自 100μL 人血浆。替格瑞洛、AR-C124910XX 和 AR-C133913XX 的批内准确度分别为 91.9-109.0%、86.8-109.2%和 100.5-112.0%；批内精密度分别为 4.0-8.4%、5.2-16.9%和 3.9-12.3%。该方法还应用于兔、大鼠、小鼠和狨猴中替格瑞洛、AR-C124910XX 和 AR-C133913XX 的定量分析，使用 25μL 动物血浆。开发了一种改良的方法来定量狗和食蟹猴血浆中的替格瑞洛和 AR-C124910XX。人源性样品结果具有良好的重现性和稳定性。该方法成功应用于人类志愿者和患者给予替格瑞洛后的血浆浓度测定以及动物安全性评价研究。该验证方法具有操作简单、稳健、高通量的优点。
