Systematic and comprehensive strategy for reducing matrix effects in LC/MS/MS analyses

化学 色谱法 液相色谱-质谱法中的离子抑制 分析物 基质(化学分析) 样品制备 固相萃取 蛋白质沉淀 电喷雾 质谱法 选择性反应监测 电喷雾电离 萃取(化学) 分析化学(期刊) 串联质谱法 生物分析 甲酸
作者
Erin E. Chambers,Diane M. Wagrowski-Diehl,Zhenhuan Lu,Jeffrey R. Mazzeo
出处
期刊:Journal of Chromatography B [Elsevier BV]
卷期号:852 (1-2): 22-34 被引量:636
标识
DOI:10.1016/j.jchromb.2006.12.030
摘要

A systematic, comprehensive strategy that optimizes sample preparation and chromatography to minimize matrix effects in bioanalytical LC/MS/MS assays was developed. Comparisons were made among several sample preparation methods, including protein precipitation (PPT), liquid-liquid extraction (LLE), pure cation exchange solid-phase extraction (SPE), reversed-phase SPE and mixed-mode SPE. The influence of mobile phase pH and gradient duration on the selectivity and sensitivity for both matrix components and basic analytes was investigated. Matrix effects and overall sensitivity and resolution between UPLC technology and HPLC were compared. The amount of specific matrix components, or class of matrix components, was measured in the sample preparation extracts by LC/MS/MS with electrospray ionization (ESI) using both precursor ion scanning mode and multiple reaction monitoring (MRM). PPT is the least effective sample preparation technique, often resulting in significant matrix effects due to the presence of many residual matrix components. Reversed-phase and pure cation exchange SPE methods resulted in cleaner extracts and reduced matrix effects compared to PPT. The cleanest extracts, however, were produced with polymeric mixed-mode SPE (both reversed-phase and ion exchange retention mechanisms). These mixed-mode sorbents dramatically reduced the levels of residual matrix components from biological samples, leading to significant reduction in matrix effects. LLE also provided clean final extracts. However, analyte recovery, particularly for polar analytes, was very low. Mobile phase pH was manipulated to alter the retention of basic compounds relative to phospholipids, whose retention tends to be relatively independent of pH. In addition to the expected resolution, speed and sensitivity benefits of UPLC technology, a paired t-test demonstrated a statistically significant improvement with respect to matrix effects when this technology was chosen over traditional HPLC. The combination of polymeric mixed-mode SPE, the appropriate mobile phase pH and UPLC technology provides significant advantages for reducing matrix effects resulting from plasma matrix components and in improving the ruggedness and sensitivity of bioanalytical methods.
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