Size-exclusion chromatography performed in capillaries. Studies by liquid chromatography-mass spectrometry.
Miniaturization of the chromatographic column led to increased sensitivity and shortened time of analysis. In our work we applied 300 microm I.D. capillaries packed with a novel stationary phase Superdex Peptide for the size-exclusion chromatography, capable of separating molecules within the mass range of 0.1-7 kDa. Here we proved that such capillary columns can operate effectively at high sensitivity. Several peptide mixtures were efficiently chromatographed and analyzed on line with electrospray ionization mass spectrometry as a detection technique. A CNBr peptide map, derived from human globin alpha subunit, was effectively separated using this method. These fragments are difficult to elute from the reversed-phase column at low pH, therefore, such approach can be considered as a complementary to other separation techniques, in particular for analyzing hydrophobic components and complex mixtures.
Smoluch
M T
MT
Faculty of Chemistry, Jagiellonian University, Krakow, Poland.
Mak
P
P
Chervet
J P
JP
Höhne
G
G
Silberring
J
J
D002850:Chromatography, Gel / Q000379:methods*; D006454:Hemoglobins / Q000737:chemistry / Q000302:isolation & purification; D006801:Humans; D013058:Mass Spectrometry / Q000379:methods*; D010446:Peptide Fragments / Q000737:chemistry / Q000302:isolation & purification; D010449:Peptide Mapping; D011957:Receptors, Opioid / Q000819:agonists; D012680:Sensitivity and Specificity
D006454:Hemoglobins; D010446:Peptide Fragments; D011957:Receptors, Opioid