Highly Sensitive Pharmaceutical and Clinical Analysis Using Selective Solid-Phase Extraction Coupled to Microflow Liquid Chromatography and Isotope-Dilution Mass Spectrometry
The capacity of ultrasensitive quantification is highly desirable in many clinical and pharmaceutical applications, such as the PK study of low-dose administration and quantification of low-abundance markers in circulating systems. Nonetheless, this task is highly challenging even for a LC-MS/MS approach, due to the extremely low levels of analytes and the highly complex matrices. Here we describe a robust method for the ultrasensitive quantification of therapeutic agents or clinical markers in highly complex biological systems. This is achieved by the combination of a selective solid-phase extraction (SPE) with a highly sensitive capillary LC (μLC)-MS/MS analysis. Comparing to a conventional LC-MS/MS, a μLC-MS/MS provides much higher sensitivity due to the lower peak dilution. SPE washing and elution conditions were optimized so that target drugs are selectively extracted from the biological matrix. By eliminating most undesirable matrix components, this selective SPE procedure enabled a high sample loading volume on the μLC column without compromising chromatographic performance and operational robustness, and helped to achieve ultralow detection limits. Sufficient μLC separation was employed in order to further improve analytical sensitivity and to decrease matrix effects. To show the application of this strategy, three paradigms, respectively, the quantifications of (1) corticosteroids after an intravitreous injection, (2) an anticancer agent after a low-dose treatment, and (3) several vitamin D (VitD) metabolites in plasma, are introduced in this chapter. Typically, low pg/mL LOQ were achieved, and the linearity, accuracy, and precision of these developed methods were excellent.
Collectively, this strategy enables the ultrasensitive, accurate, and robust LC-MS-based quantification of extremely low levels of drug/markers in biological samples, which were previously approachable only by ELISA or RIA methods.
KeywordsTarget Compound Collisionally Activate Dissociation Mobile Phase Modifier Complex Biological Matrice Separation Segment
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