用于定量测定血浆、尿液、粪便、肝脏和肾脏中伊立替康、SN-38和SN-38葡萄糖醛酸苷的超高效液相色谱-串联质谱法的开发与验证：在大鼠伊立替康药代动力学研究中的应用
Development and validation of an UPLC-MS/MS method for the quantification of irinotecan, SN-38 and SN-38 glucuronide in plasma, urine, feces, liver and kidney: Application to a pharmacokinetic study of irinotecan in rats.
作者信息
Basu Sumit, Zeng Min, Yin Taijun, Gao Song, Hu Ming
机构信息
Department of Pharmacological and Pharmaceutical Sciences, College of Pharmacy, University of Houston, 1441 Moursund Street, Houston, TX 77030, USA; Center for Pharmacometrics and Systems Pharmacology, College of Pharmacy, University of Florida, 6550 Sanger Road, Orlando, FL 32827, USA.
Department of Pharmacological and Pharmaceutical Sciences, College of Pharmacy, University of Houston, 1441 Moursund Street, Houston, TX 77030, USA; Department of Thoracic and Cardiomacrovascular surgery, Shiyan Taihe Hospital Affiliated to Hubei University of Medicine, Shiyan, Hubei 442000, China.
出版信息
J Chromatogr B Analyt Technol Biomed Life Sci. 2016 Mar 15;1015-1016:34-41. doi: 10.1016/j.jchromb.2016.02.012. Epub 2016 Feb 9.
The objective of this research is to develop and validate a sensitive and reproducible UPLC-MS/MS method to quantify irinotecan, its active metabolite SN-38 and SN-38 glucuronide (phase II metabolite of SN-38) simultaneously in different bio-matrices (plasma, urine, feces), tissues (liver and kidney) and to use the method to investigate its pharmacokinetic behavior in rats. Irinotecan, SN-38 and SN-38 glucuronide has been resolved and separated by C18 column using acetonitrile and 0.1% formic acid in water used as the mobile phases. Triple quadruple mass spectrometer using multiple reaction monitoring (MRM) with positive scan mode were employed to perform mass analysis. The results showed that the linear response range of irinotecan and SN-38 in plasma, feces, liver and kidney is 4.88-10000 nM, 39-5000 nM, 48.8-6250 nM and 48.8-6250 nM, respectively (R(2)>0.99). In case of SN-38 glucuronide, the standard curves were linear in the concentration range of 6.25-2000 nM, 4.88-1250 nM, 9.8-1250 nM and 9.8-1250 nM in plasma, feces, liver and kidney homogenates, respectively. The lower limit of detection (LLOD) of irinotecan, SN-38 and SN-38 glucuronide was determined to be less than 25 nM in all bio-matrices as well as tissue homogenates. Recoveries of irinotecan, SN-38 and SN-38 glucuronide at three different concentrations (low, medium and high) were not less than 85% at three different concentrations in plasma and feces. The percentage matrix factors in different bio-matrices and tissues were within 20%. The UPLC-MS/MS method was validated with intra-day and inter-day precision of less than 15% in plasma, feces, liver and kidney. Owing to the high sensitivity of this method, only 20 μl of plasma, urine and homogenates of liver, kidney and feces is needed. The validated method has been successfully employed for pharmacokinetic evaluation of irinotecan in male wistar rats to quantify irinotecan, SN-38 and SN-38 glucuronide in plasma, feces, and urine samples.
本研究的目的是开发并验证一种灵敏且可重复的超高效液相色谱-串联质谱法，用于同时定量不同生物基质（血浆、尿液、粪便）、组织（肝脏和肾脏）中的伊立替康及其活性代谢物SN-38和SN-38葡萄糖醛酸苷（SN-38的Ⅱ相代谢物），并使用该方法研究其在大鼠体内的药代动力学行为。伊立替康、SN-38和SN-38葡萄糖醛酸苷已通过C18柱进行分离，流动相采用乙腈和0.1%甲酸水溶液。采用三重四极杆质谱仪，在正离子扫描模式下通过多反应监测（MRM）进行质谱分析。结果表明，伊立替康和SN-38在血浆、粪便、肝脏和肾脏中的线性响应范围分别为4.88 - 10000 nM、39 - 5000 nM、48.8 - 6250 nM和48.8 - 6250 nM（R(2)>0.99）。对于SN-38葡萄糖醛酸苷，其在血浆、粪便、肝脏和肾脏匀浆中的标准曲线在浓度范围6.25 - 2000 nM、4.88 - 1250 nM、9.8 - 1250 nM和9.8 - 1250 nM内呈线性。在所有生物基质以及组织匀浆中，伊立替康、SN-38和SN-38葡萄糖醛酸苷的检测下限（LLOD）均测定为小于25 nM。伊立替康、SN-38和SN-38葡萄糖醛酸苷在血浆和粪便中三个不同浓度（低、中、高）的回收率在三个不同浓度下均不低于85%。不同生物基质和组织中的基质因子百分比在20%以内。该超高效液相色谱-串联质谱法在血浆、粪便、肝脏和肾脏中的日内和日间精密度均小于15%，从而得到验证。由于该方法灵敏度高，仅需20 μl血浆、尿液以及肝脏、肾脏和粪便的匀浆。已验证的该方法已成功用于雄性Wistar大鼠伊立替康的药代动力学评价，以定量血浆、粪便和尿液样本中的伊立替康、SN-38和SN-38葡萄糖醛酸苷。
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