Anal. Chem. 1997, 69, 51-60
Capillary Zone Electrophoresis-Electrospray Mass Spectrometry at Submicroliter Flow Rates: Practical Considerations and Analytical Performance John F. Kelly,†,‡ Louis Ramaley,† and Pierre Thibault*,§
Department of Chemistry, Dalhousie University, Halifax, Nova Scotia, and Institute for Marine Biosciences, National Research Council Canada, 1411 Oxford Street, Halifax, Nova Scotia, B3H 3Z1, Canada
A novel method for the production of coated capillaries for capillary zone electrophoresis-electrospray mass spectrometry (CZE-ESMS), operating at 100-200 nL/min flow rates is described. Electrophoretic conditions conducive to the simultaneous operation of ESMS were investigated for microsprayers of different inner diameters. The ruggedness and reliability of the CZE columns with microsprayers of 20 µm i.d. were investigated over 6 h of uninterrupted operation. Detection limits in the order of 0.1-5 fmol were obtained for peptide standards of molecular masses ranging from 500 to 2500 Da, and sensitivity was ∼10 times higher than that achieved using the more conventional sheath flow CZE-ESMS interface. Such enhancement of signal detectability provided distinct advantages for on-line line tandem mass spectrometric analyses whereby structurally informative MS-MS spectra were obtained using femtomole loadings of peptides and glycopeptides from ovalbumin digests. The combination of capillary zone electrophoresis (CZE) with electrospray mass spectrometry (ESMS) provides a powerful tool for the separation and characterization of charged analytes and biomolecules at sub-picomole levels. Since the first report of CZE-ESMS in 1987 by Olivares et al.,1 this technique has undergone significant instrumental developments which have been parallelled by an impressive number of applications, ranging from the separation of small inorganic ions2,3 to the analysis of complex biomolecules such as peptides and proteins.4-10 A description of these applications can be found in a number of excellent reviews †
Dalhousie University. Current address: Merck Frosst, Point Claire, PQ, Canada. § Institute for Marine Biosciences. (1) Olivares, J. A.; Nguyen, N. T.; Yonker, C. R.; Smith, R. D. Anal. Chem. 1987, 59, 1230-1232. (2) Huggins, T. G.; Henion, J. D. Electrophoresis 1993, 14, 531-539. (3) Corr, J. J.; Covey, T. R.; Anacleto, J. F. Proceedings, 42nd ASMS Conference on Mass Spectrometry and Allied Topics, Chicago, IL, 29 May-3 June 1994; pp 340. (4) Loo, J. A.; Udseth, H. R.; Smith, R. D. Anal. Biochem. 1989, 179, 404-412. (5) Moseley, M. A.; Jorgenson, J. J.; Shabanowitz, J.; Hunt, D. F.; Tomer, K. B. J. Am. Soc. Mass Spectrom, 1992, 3, 289-300. (6) Smith, R. D.; Udseth, H. R.; Barinaga, C. J.; Edmonds, C. G. J. Chromatogr. 1991, 559, 197-208. (7) Lee, E. D.; Muck, W.; Henion, J. D.; Covey, T. R. J. Chromatogr. 1989, 458, 313. (8) Tsuji, K.; Baczynsky, L.; Bronson, G. E. Anal. Chem. 1992, 64, 1864-1870. (9) Thibault, P.; Paris, C.; Pleasance, S. Rapid Commun. Mass Spectrom. 1991, 542, 483-501. (10) Thompson, T.; Foret, F.; Vouros, P.; Karge, B. L. Anal. Chem. 1993, 65, 900-906. ‡
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© 1996 American Chemical Society
on CZE-ESMS that have appeared in the literature over the past five years.11-15 The on-line coupling of CZE to ESMS has been particularly challenging in view of the low flow rates typically achievable under zone electrophoresis (
