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Development of an LC–MS method to quantify coproporphyrin I and III as endogenous biomarkers for drug transporter-mediated drug-drug interactions
Journal of Chromatography B ( IF 3 ) Pub Date : 2017-12-06 , DOI: 10.1016/j.jchromb.2017.12.008
Emmanuel Njumbe Ediage , Lieve Dillen , Ann Vroman , Luc Diels , Annett Kunze , Jan Snoeys , Tom Verhaeghe

Coproporphyrins are proposed as endogenous biomarkers of hepatic Organic Anion Transporting Polypeptide (OATP)1 B functional activity. In this study, a new sample extraction method based on a mixed-mode anion exchange sorbent (SPE clean-up using Oasis 30 mg Max 96 well plates) was developed for absolute quantification of coproporphyrin I and III (CP-I and CP-III) in human plasma. Chromatographic separation was performed with an Ace Excel 2 C18 PFP, 3 μm, 2.1 × 150 mm, maintained at 60 °C. A 10 mM ammonium formate containing 0.1% HCOOH and acetonitrile (100%) was used as mobile phase A and B, respectively. Mass transition, m/z 655.3 → 596.3 was selected to monitor CP-I and CP-III, while m/z 659.3 → 600.3 transition was used for the stable isotope labelled internal standard. Optimization of the liquid chromatography tandem mass spectrometry method ensured a lower limit of quantification (LLOQ) of 20 pg/mL. Both CP-I and CP-III had extraction recoveries of 70%. The calibration range was 0.02–100 ng/mL for both CP-I and CP-III, yielding calibration curves with correlation coefficients greater than 0.988. Inter day precision (CV < 9%) and accuracy (84.3–103.9%) complied with the recommendation of the European Bioanalytical Forum.

The optimized method was used to analyse plasma samples originating from three independent clinical studies. Obtained CP-I and CP-III plasma baseline levels in healthy volunteers were in good agreement with previously published data. Moreover, CP-I and CP-III plasma levels in human subjects dosed with a clinically confirmed OATP inhibitor were significantly increased compared to their baseline levels. These data demonstrate the potential of CP-I and CP-III as endogenous biomarkers to predict the drug-drug interaction (DDI) related to hepatic OATP1 B inhibition. Stability of CP-I and CP-III in plasma and solvents under different processing and storage conditions was also evaluated.



中文翻译:

LC-MS方法的开发,以定量定量作为药物转运蛋白介导的药物相互作用的内源性生物标志物的卟啉I和III

拟卟啉被认为是肝有机阴离子转运多肽(OATP)1 B功能活性的内源性生物标志物。在这项研究中,开发了一种基于混合模式阴离子交换吸附剂(使用Oasis 30 mg Max 96孔板净化SPE)的新样品萃取方法,用于对卟啉I和III(CP-I和CP-III)进行绝对定量)在人体血浆中。色谱分离是使用Ace Excel 2 C18 PFP,3μm,2.1×150 mm,保持在60°C进行的。分别将含有0.1%HCOOH和10%乙腈(100%)的10 mM甲酸铵用作流动相A和B。质量转换m / z选择655.3→596.3来监视CP-I和CP-III,而m / z659.3→600.3过渡用于稳定同位素标记的内标。液相色谱串联质谱法的优化确保了20 pg / mL的定量下限(LLOQ)。CP-I和CP-III的提取回收率均为70%。CP-I和CP-III的校准范围均为0.02–100 ng / mL,所产生的校准曲线的相关系数大于0.988。日间精度(CV <9%)和精度(84.3–103.9%)符合欧洲生物分析论坛的建议。

优化的方法用于分析源自三个独立临床研究的血浆样品。在健康志愿者中获得的CP-I和CP-III血浆基线水平与先前发表的数据高度吻合。而且,与基线水平相比,在临床上确认为OATP抑制剂的人类受试者中CP-1和CP-III血浆水平显着增加。这些数据表明,CP-1和CP-III作为内源性生物标志物的潜力,可预测与肝OATP1 B抑制有关的药物相互作用。还评估了CP-1和CP-3在不同处理和储存条件下在血浆和溶剂中的稳定性。

更新日期:2017-12-06
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