Comparison between transient isotachophoretic capillary zone electrophoresis and reversed-phase liquid chromatography for the determination of peptides in plasma

Low levels of peptide drugs in human plasma can be determined employing off -line solid-phase extraction, followed by capillary zone electrophoresis wi th UV detection. A bioanalytical procedure is presented, using gonadorelin and angiotensin II in human plasma as model compounds. The solid-phase extr action method, based on a weak cation exchange mechanism, is able to remove interfering endogenous components from the plasma sample, extract the mode l peptides quantitatively, and give a possibility of concentrating the samp le at the same time. Transient isotachophoretic conditions were kept to inc rease the sample loadability by about two orders of magnitude. Up to about 70% of the capillary was filled with the reconstituted extract, whereafter the peptides were selectively concentrated during the first 15 min. Subsequ ently, the concentrated sample zones were separated under capillary zone el ectrophoresis conditions, showing the technique's high resolution. For the model cationic peptides (gonadorelin, angiotensin II) good linearity and re producibility was observed in the 20-100 ng/mL concentration range. A more extensive washing procedure permits quantitation of gonadorelin at the 5 ng /mL level. In comparison with a liquid chromatography analysis, superior ma ss sensitivity and separation are obtained with the transient isotachophore tic capillary zone electrophoresis method. Moreover, in this case equivalen t sensitivity is achieved when it is directly compared with a liquid chroma tography method with UV detection, keeping in mind that 60 times more sampl e is needed for the latter method. A further gain in sensitivity can be obt ained when the analysis is combined with native fluorescence detection, as is demonstrated by combining liquid chromatography separation with fluoresc ence detection.