Chromatographic preconcentration coupled to capillary electrophoresis via an in-line injection valve.
A preconcentration-capillary electrophoresis (CE) system using a small precolumn in combination with an in-line injection valve is presented. The advantage of the present design is the ability to perform the sample preconcentration fully independently from the CE separation and to prevent sample matrix and washing solvents from entering the CE capillary. With a micro injection valve, sample could be effectively introduced into the CE system in an in-line fashion without seriously affecting the CE separation efficiency. Breakthrough volume, desorption efficiency, and elution volume for the C18 microcolumn (5 x 0.5 mm i.d.) were established, yielding values of 750 microL, 70%, and 0.9-1.1 microL, respectively, using enkephalin peptides. The time between the start of the desorption of the analytes from the precolumn and the injection into the CE system was also studied in order to achieve optimal sensitivity and separation efficiency. The performance of the complete system was demonstrated by the preconcentration and separation of an enkephalin mixture. Using a sample volume of 250 microL and a CE injection voltage of -15 kV for 12 s, linearity was observed over 2 orders of magnitude, and detection limits (S/N = 3) were in the 5-10 ng/mL range. A 1000-fold sensitivity enhancement is obtained using this setup, as compared to a regular CE setup. For 100 ng/mL samples, repeatabilities (RSDs) of migration time and peak area were 1.2 and 11%, respectively.
Tempels
F W Alexander
FW
Department of Biomedical Analysis, Faculty of Pharmaceutical Sciences, Utrecht University, Sorbonnelaan 16, NL-3584 CA Utrecht, The Netherlands.
Underberg
Willy J M
WJ
Somsen
Govert W
GW
de Jong
Gerhardus J
GJ
D002845:Chromatography / Q000295:instrumentation / Q000379:methods; D019075:Electrophoresis, Capillary / Q000295:instrumentation / Q000379:methods*; D004745:Enkephalins / Q000737:chemistry* / Q000302:isolation & purification; D004867:Equipment Design; D007202:Indicators and Reagents; D010446:Peptide Fragments / Q000737:chemistry / Q000302:isolation & purification; D012997:Solvents
D004745:Enkephalins; D007202:Indicators and Reagents; D010446:Peptide Fragments; D012997:Solvents