Journal
JOURNAL OF CHROMATOGRAPHY B-ANALYTICAL TECHNOLOGIES IN THE BIOMEDICAL AND LIFE SCIENCES
Volume 853, Issue 1-2, Pages 234-241Publisher
ELSEVIER SCIENCE BV
DOI: 10.1016/j.jchromb.2007.03.017
Keywords
liquid-liquid extraction; pH adjustment; bioanalysis; modelling; recovery; selectivity
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Liquid-liquid extraction (LLE) is widely used as a simple and robust sample preparation technique in bioanalytical sample preparation. When extracting ionisable compounds, most bioanalysts adjust the pH of the sample to achieve fully unionized compounds. Usually, a generally accepted rule is applied to adjust the pH of the aqueous phase, known as the pK(a) +/- 2 rule, depending on the acid/base characteristics of the analyte. By taking a closer look at the general equations that describe the extraction behaviour of ionisable compounds, we extended this pH adjustment rule by taking the distribution ratio and the volume of both liquid phases into account. By choosing an extraction pH based on this extended rule, the selectivity of the extraction can be influenced without loss of recovery. As a measure of this selectivity, two equations were proposed to indicate the ability of the extraction system to discriminate between two compounds. Also, milder extraction pH can be used for pH labile analytes. To use this new rule quantitatively, a new calculation method for the determination of the distribution ratio was derived. These calculations were based on normalized recoveries making this method less susceptible to errors in absolute recovery determination. The proposed equations were supported by demonstrating that careful pH adjustment can lead to higher selectivity. The main conclusion was that a closer look at the extraction pH in bioanalytical methods extends the possibilities of obtaining a higher selectivity or the possibilities of extracting pH labile analytes at milder pH conditions without loss of recovery. (c) 2007 Elsevier B.V. All rights reserved.
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