Development of an ultra fast online-solid phase extraction (SPE) liquid chromatography electrospray tandem mass spectrometry (LC-ESI-MS/MS) based approach for the determination of drugs in pharmacokinetic studies
Development of an ultra fast online-solid phase extraction (SPE) liquid chromatography electrospray tandem mass spectrometry (LC-ESI-MS/MS) based approach for the determination of drugs in pharmacokinetic studies
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DOI：
10.1039/c0ay00714e
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发表时间：
2011-02-01
影响因子：
3.1
通讯作者：
Hammock, Bruce D.
中科院分区：
文献类型：
--
作者：
Schebb, Nils Helge;Inceoglu, Bora;Hammock, Bruce D.
药物开发过程中，对大量药代动力学（PK）研究样本进行高通量分析至关重要。使用LC-ESI-MS分析来自血液的候选药物通常需要分离血浆级分，然后进行各种离线样品制备程序。此步骤是阻碍吞吐量的瓶颈。为了克服这一困难并加速PK和其他研究中的分析，我们开发了一种方法，允许在稀释和离心后直接分析低体积的全血（10 μ L）。样品在在线-SPE-LC-ESI-MS/MS设置中进样，使得完全梯度分离的总运行时间仅为126 s。通过湍流色谱法在30 s内从基质中提取分析物。随后，在50 × 2.1 mm（1.7 μ m）RP-18色谱柱上在1.5分钟内进行完全梯度分离，并在SRM模式下通过ESI-MS/MS灵敏地检测分析物。这种新的超快速在线SPE-LC-ESI-MS/MS方法的性能通过分析广泛使用的抗炎药物双氯芬酸（DCF）得到了证明。DCF以稳定的保留时间（+/-0.33%）洗脱，峰宽窄（FWHM 3.3 +/-0.15 s）。该方法显示出优异的分析性能，检测限为6 fmol的柱上，线性范围超过四个数量级和一个可忽略不计的携带0.12 +/- 0.03%的DCF。使用该方法研究了大鼠局部和腹膜内给药以及1名志愿者经口给药DCF的PK特征。最后，通过分析罗非昔布和几种可溶性环氧化物水解酶抑制剂，证明了该方法对药物的普遍适用性。这种新方法只需要现成的标准LC仪器，并符合绿色分析化学的要求。
High-throughput analyses of a large number of samples for pharmacokinetic (PK) studies are essential in drug development. Analysis of drug candidates from blood using LC-ESI-MS generally requires separation of the plasma fraction followed by various offline sample preparation procedures. This step is a bottleneck that impedes throughput. In order to overcome this difficulty and accelerate analysis in PK and other studies, we developed an approach allowing the direct analysis of low volumes of whole blood (10 mu L) after dilution and centrifugation. Samples were injected in an online-SPE-LC-ESI-MS/MS setup allowing a total run time of only 126 s for a full gradient separation. Analytes were extracted from the matrix within 30 s by turbulent flow chromatography. Subsequently, a full gradient separation was carried out within 1.5 minutes on a 50 x 2.1 mm (1.7 mu m) RP-18 column and the analytes were sensitively detected by ESI-MS/MS in SRM mode. The performance of this new ultra fast online SPE-LC-ESI-MS/MS approach was demonstrated by the analysis of diclofenac (DCF), a widely used anti-inflammatory drug. DCF eluted at stable retention times (+/-0.33%) with narrow peak width (FWHM 3.3 +/- 0.15 s). The method displays excellent analytical performance, with a limit of detection of 6 fmol on column, a linear range of over four orders of magnitude and a negligible carry over of 0.12 +/- 0.03% for DCF. The PK profile of DCF administered by topical and intraperitoneal routes in rats and by oral route in one human volunteer is investigated using this method. Finally, general applicability of the approach for drugs is demonstrated by analysis of rofecoxib and several inhibitors of the soluble epoxide hydrolase. This new method requires only readily available, off the shelf standard LC instrumentation, and is compliant with the requirements of green analytical chemistry.