Analysis of omeprazole and 5-OH omeprazole in human plasma using hydrophilic interaction chromatography with tandem mass spectrometry (HILIC-MS/MS) - Eliminating evaporation and reconstitution steps in 96-well liquid/liquid extraction
Analysis of omeprazole and 5-OH omeprazole in human plasma using hydrophilic interaction chromatography with tandem mass spectrometry (HILIC-MS/MS) - Eliminating evaporation and reconstitution steps in 96-well liquid/liquid extraction
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DOI：
10.1016/j.jchromb.2005.10.042
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发表时间：
2006-01-02
影响因子：
3
通讯作者：
Weng, ND
中科院分区：
文献类型：
--
作者：
Qi, S;Weng, ND
使用液/液萃取（LLE）和液相色谱-电喷雾串联质谱（LC-MS/MS）的生物分析方法被广泛使用。有机提取物需要蒸发和重构，阻碍了生产量和自动化的进一步提高。在这项研究中，我们证明了一种新的方法，消除这两个步骤，在96孔LLE通过使用亲水相互作用色谱与MS/MS（HILIC-MS/MS）的硅胶柱上的高有机/低水移动的流动相。在96孔板中，使用0.5 ml乙酸乙酯从0.05 ml人血浆中提取奥美拉唑、其代谢产物5-OH奥美拉唑和内标物脱氧奥美拉唑。用0.4 ml乙腈稀释一部分（0.1 ml）乙酸乙酯提取物，并将10 μ l注入Betasil硅胶柱（50 mm x 3.0 mm，5 μ m）中，用API 3000和4000（+）ESI检测。具有线性梯度洗脱的移动的相由乙腈、水和甲酸组成（在2分钟内从95：5：0.1至73.5：26.5：0.1）。流速为1.5 ml/min，总运行时间为2.75 min。该方法经验证，两种分析物的定量下限均为2.5 ng/ml。还对该方法的专属性、重现性、稳定性和回收率进行了验证。还证明了无不良基质效应和残留。低、中、高浓度水平质控样品的日间精密度和准确度均< 4.4%相对标准偏差（R.S.D.）和4.1%的相对误差（R.E.）奥美拉唑的相对标准偏差为4.5%。和5.6%的R. 5-OH奥美拉唑。(c)2005 Elsevier B. V.保留所有权利。
Bioanalytical methods using liquid/liquid extraction (LLE) and liquid chromatography with electrospray tandem mass spectrometry (LC-MS/MS) are widely used. The organic extracts need to be evaporated and reconstituted, hampering further improvement of throughput and automation. In this study, we demonstrated a novel approach of eliminating these two steps in 96-well LLE by using hydrophilic interaction chromatography with MS/MS (HILIC-MS/MS) on silica column with high organic/low aqueous mobile phase. Omeprazole, its metabolite 5-OH omeprazole, and internal standard desoxyomeprazole, were extracted from 0.05 ml of human plasma using 0.5 ml of ethyl acetate in a 96-well plate. A portion (0.1 ml) of the ethyl acetate extract was diluted with 0.4 ml of acetonitrile and 10 mu l was injected onto a Betasil silica column (50 mm x 3.0 mm, 5 mu m) and detected by API 3000 and 4000 with (+) ESI. Mobile phase with linear gradient elution consists of acetonitrile, water, and formic acid (from 95:5:0.1 to 73.5:26.5:0.1 in 2 min). The flow rate was 1.5 ml/min with total run time of 2.75 min. The method was validated for a low limit of quantitation at 2.5 ng/ml for both analytes. The method was also validated for specificity, reproducibility, stability and recovery. Lack of adverse matrix effect and carry-over was also demonstrated. The inter-day precision and accuracy of the quality control samples at low, medium and high concentration levels were < 4.4% relative standard deviation (R.S.D.) and 4.1% relative error (R.E.) for omeprazole, and 4.5% R.S.D. and 5.6% R.E. for 5-OH omeprazole, respectively. (c) 2005 Elsevier B.V. All rights reserved.