Quantitation of efletirizine in human plasma and urine using automated solid-phase extraction and column-switching high-performance liquid chromatography

A heart-cut column-switching, ion-pair, reversed-phase HPLC system was used for the quantitation of efletirizine (EFZ) in biological fluids. The analyte and an internal standard (I.S.) were extracted from human EDTA plasma by C18 solid-phase extraction (SPE) using a RapidTrace workstation. The eluent from the SPE was evaporated, reconstituted and injected onto the HPLC column. Urine samples were diluted and injected directly without the need of extraction. The compounds of interest were separated from most of the extraneous matrix Materials by the first C18 column, and switched onto a second C18 column for further separation using a mobile phase of stronger eluting capability. Linearity range was 10-2000 ng ml(-1) for plasma and 0.05-10 microg ml(-1) for urine. The lower limit of quantitation (LOQ) was 10 ng from 1 ml of plasma, with a signal-to-noise ratio of 15:1. Inter-day precision and bias of quality control samples (QCs) were <5% for plasma and <7% for urine. Selectivity was established against six other antihistamines, three analogs of efletirizine, and on 12 control plasma lots and nine control urine lots. Recovery was 90.0% for EFZ and 89.5% for I.S. from plasma. One hundred samples can be processed in every 2.75 h on a 10-module RapidTrace workstation with minimal human attention. Method ruggedness were tested on three brands of SPE and six different lots of one SPE brand. Performance ruggedness was demonstrated by different analysts on multiple HPLC systems. Analyte stability through sample storage, extraction process (benchtop, freeze-thaw, refrigeration after extraction) and chromatography (on-system, reinjection) was established.