Separation of nanoliter samples of biological amines by a comprehensive two-dimensional microcolumn liquid chromatography system.
A two-dimensional liquid chromatography system based on the combination of a charge separation mechanism, provided by anion exchange chromatography, with a hydrophobic separation mechanism, provided by reversed phase chromatography, is presented. A 90 cm long anion exchange microcolumn is coupled to a 3 cm long reversed phase microcolumn. Both microcolumns are interfaced by two electronically controlled valves. Effluent from the anion exchange microcolumn is collected in a sample loop and then concentrated onto the head of the reversed phase microcolumn. Analyte peaks consisting of tagged amines elute from the second column and are detected by laser-induced fluorescence. The entire operation is controlled by a personal computer. The resolving power of this system is demonstrated with a two-dimensional chromatogram of the peptides obtained from a tryptic digest of porcine thyroglobulin. The sensitivity of the system is demonstrated with a two-dimensional chromatogram of the contents of a single bovine chromaffin cell.
Holland
L A
LA
Department of Chemistry, University of North Carolina at Chapel Hill 27599-3290, USA.
Jorgenson
J W
JW
D000588:Amines / Q000302:isolation & purification*; D000818:Animals; D002417:Cattle; D002838:Chromaffin System / Q000737:chemistry / Q000166:cytology; D002852:Chromatography, Ion Exchange; D013552:Swine; D013954:Thyroglobulin / Q000032:analysis
D000588:Amines; D013954:Thyroglobulin