Negligible Depletion Solid-Phase Microextraction with Radiolabeled Analytes To Study Free Concentrations and Protein Binding: An Example with [3H]Estradiol

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• 作者：Minne B. Heringa ; Dolors Pastor ; Jon Algra ; Wouter H. J. Vaes ; and Joop L. M. Hermens
• 刊名：Analytical Chemistry
• 出版年：2002
• 出版时间：December 1, 2002
• 年：2002
• 卷：74
• 期：23
• 页码：5993 - 5997
• 全文大小：61K
• 年卷期：v.74,no.23(December 1, 2002)
• ISSN：1520-6882

文摘

A new method is presented that enables sensitive measurement of free concentrations of radiolabeled ligands.Additionally, protein binding of radiochemicals in complex matrixes can be determined with this new techniquethat combines negligible depletion solid-phase microextraction (nd-SPME) with liquid scintillation counting(LSC) as detection. [³H]Estradiol was taken as an examplecompound. Possible matrix effects of protein on fiberuptake kinetics were studied. No matrix effect was found,either by fouling of the fiber, or by changed uptakekinetics. The validity of the method was shown in thedetermination of the affinity constant (K_a) of estradiol forhuman serum albumin (HSA). The K_a was estimated at8.9 × 10⁴ M^-1, which corresponds well with literaturevalues. This study shows that nd-SPME is suitable tostudy the free concentration and protein binding of [³H]estradiol. The method described in this paper combinesthe advantages of nd-SPME with the advantages of radiolabeled analytes, creating a timesaving, simple, and sensitive analytical tool that will be particularly useful incomplex matrixes containing many potential interferencesfor chromatographic methods.