On-line preconcentration capillary electrophoresis for purity profiling of synthetic peptides

VIZIOLI NM*; RUSELL ML; CARDUCCI CN

ANALYTICA CHIMICA ACTA

Elsevier

Lugar: New York; Año: 2004 vol. 514 p. 167 - 177

0003-2670

In this work, an on-line preconcentration capillary electrophoresis method was optimized and evaluated for the purity control of the biologically active synthetic peptide fas-F (a 28-residue long fragment of fasciculin-1) and applied for the purity profiling of angiotensin I, oxytocin, bradykinin and luteinizing hormone releasing hormone. The laboratory-made device of the analyte concentrator cartridge consisted of a fused-silica capillary piece (150 ¦Ìm i.d.¡Á8 mm in length) packed with silica-based C18 reversed-phase chromatographic material and coupled on-line near the inlet of the separation capillary (bare fused-silica capillary, 75 ¦Ìm i.d.¡Á28 cm from the concentrator to the detector). Separation of impurities present in assayed samples was achieved by using 25 mM potassium dihydrogen phosphate, pH 3.5, as running buffer and a mixture of acetonitrile: running buffer, 75:25 (v/v), as elution buffer. The intra-day relative standard deviation (R.S.D.) values for migration times ranged from 3.4 to 4.2 and 2.2¨C2.6% for peak areas. The inter-day R.S.D. values were 5.6¨C7.1 and 4.6¨C5.1% for migration times and peak areas, respectively. The impurity profiles obtained for each peptide were compared by CZE and on-line preconcentration CE. The proposed method allowed the preconcentration and separation of impurities with greater selectivity and higher sensitivity (100¨C200-fold) with respect to capillary zone electrophoresis without on-line preconcentration.