A fully automated on-line salting-out assisted liquid-liquid extraction capillary electrophoresis methodology: Application to tyrosine kinase inhibitors in human plasma

Abstract

A fully automated analytical methodology combining salting-out assisted liquid-liquid extraction (SALLE) and capillary electrophoresis (CE) for the analysis of three Tyrosine Kinase Inhibitors (TKIs) in plasma samples is proposed. The automated methodology, called A-SALLE-CE-UV, makes full use of the advantages of both techniques by combining desalting, protein precipitation, automated liquid-liquid extraction, in-line CE stacking and electrophoretic separation of analytes in plasma samples in a fully integrated way. At first, the capillary is used to deliver appropriate micro-volumes of extraction agent solutions (acetonitrile, salt) in the plasma sample. ACN and salting-out agent (NaCl) solutions are added by pressure from outlet vials into the sample vial (inlet) containing human plasma sample spiked with the three tested TKIs. After addition of both ACN and NaCl solutions, mixing is achieved by generating air bubbles leading to a two phases separation and extraction of TKIs in the upper mostly organic phase (ACN). The upper phase containing the TKIs is then injected and analysed by CE-UV. Due to the presence of ACN, the analytes are stacked in-line and successfully separated in the same capillary. The results obtained in terms of limit of detection (LOD), limit of quantification (LOQ), sensitivity enhancement factor (SEF), repeatability and linearity demonstrate the applicability of the proposed method for possible therapeutic drug monitoring (TDM) of TKIs.