摘要:
液相色谱-串联质谱(LC-MS/MS)法具有高灵敏度、高选择性、高通量等特点,已经成为生物分析的主流方法,广泛应用于新药发现和开发过程中新化学实体及其代谢物的定量分析.然而,由于生物样品基质成分复杂,共洗脱物质会影响分析物的离子化,使分析物的质谱响应增加或降低,从而影响LC-MS/MS分析方法的准确度和精密度.一般而言,离子抑制较离子增强更为常见.因此,在建立LC-MS/MS法时,需要对离子抑制进行评估和校正,并采用不同的策略消除或减少基质效应的影响.本文综述了生物样品分析中基质效应的来源和评估方法,重点介绍了克服基质效应的策略.引起基质效应的物质包括磷脂、盐类、尿素、代谢物等内源性物质和赋形剂、抗凝剂、固定相释放物质及降解产物等外源性物质.目前基质效应的评价方法主要有提取后加入法和柱后灌注法.克服基质效应的策略主要有使用稳定同位素内标,将极性药物衍生化,沉淀蛋白后稀释上清液,优化样品预处理方法、色谱和质谱条件等.结合本课题组的应用实例,重点阐述了建立LC-MS/MS生物分析方法时,为减少基质效应遇到的困难及解决方法.
Abstract:
Liquid chromatography-tandem mass spectrometry (LC-MS/MS) method has the characteristics of high sensitivity, high selectivity and high throughput, and has become the mainstream approach for bioanalytical assays.It is widely used for the quantitative analysis of new drug entities (NCEs) and their metabolites in the discovery and development of new drugs.However, due to the complex composition of the biological sample matrix, the co-eluting material affects the ionization of the analyte, result in increases or decreases the mass spectral response of the analyte, thus affecting the accuracy and precision of the LC-MS/MS analysis method.In general, ion suppression is more common than ion enhancement.When the analytical method is established, if the ion suppression is not evaluated and corrected, the sensitivity of the analytical method will be seriously affected.Therefore, it is important to assess the matrix effect and use different strategies to eliminate or reduce the impact of matrix effects.In this paper, an overview was provided, including how matrix effect occurs, outline the methodologies used to assess the matrix effect in biological sample analysis and discuss the strategy of overcoming the matrix effect.Substances that cause matrix effects include endogenous substances and exogenous substances.Endogenous substances include phospholipid, salts, urea, metabolites and so on.Exogenous substances such as excipients, anticoagulants, stationary phase release materials and degradation products.At present, there are mainly two method to assess the matrix effect, one is post-extraction spike, the other is post-column infusion.The strategies to overcome the matrix effect including the use of stable-isotope labeled internal standard, derivatization of polar drugs, dilute the supernatant after precipitation of the protein, optimize sample pretreatment method, the chromatographic conditions and the mass spectrometry conditions.This paper provides examples of the application of our laboratory, focusing on the difficulties encountered and solutions in the establishment of LC-MS/MS analysis methods.
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