Capillary electrophoresis-mass spectrometry of proteins at medium pH using bilayer-coated capillaries

The feasibility of using noncovalently bilayer-coated capillaries for capillary electrophoresis-mass spectrometry (CE-MS) of acidic proteins was investigated using background electrolytes (BGEs) of medium pH. The capillary was coated by successively rinsing the capillary with solutions of the opposi...

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Bibliographic Details
Published in:Analyst (London) 2007-01, Vol.132 (1), p.75-81
Main Authors: CATAI, Jonatan R, TORANO, Javier Sastre, DE JONG, Gerhardus J, SOMSEN, Govert W
Format: Article
Language:English
Subjects:
Analytical chemistry
Animals
Chemistry
Chromatographic methods and physical methods associated with chromatography
Electrophoresis, Capillary - instrumentation
Electrophoresis, Capillary - methods
Exact sciences and technology
Human Growth Hormone - analysis
Humans
Hydrogen-Ion Concentration
Other chromatographic methods
Proteins - analysis
Spectrometric and optical methods
Spectrum Analysis - methods
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Summary:The feasibility of using noncovalently bilayer-coated capillaries for capillary electrophoresis-mass spectrometry (CE-MS) of acidic proteins was investigated using background electrolytes (BGEs) of medium pH. The capillary was coated by successively rinsing the capillary with solutions of the oppositely charged polymers polybrene (PB) and poly(vinyl sulfonic acid) (PVS). Volatile BGEs containing ammonium formate and/or N-methyl morpholine were tested at pH 7.5 and 8.5. Overall, these BGEs provided relatively fast protein separations (analysis times of ca. 12 min) and showed high efficiencies (70,000-300,000 plates) when the ionic strength was sufficiently high. Migration-time reproducibilities were very favorable with RSDs of less than 1.0%. Infusion experiments showed satisfactory MS responses for studied proteins dissolved in ammonium formate (pH 8.5), however, high concentrations of N-methyl morpholine appeared to seriously suppress the MS protein signals. Evaluation of the CE-MS system was performed by analyzing a mixture of intact proteins yielding efficient separations and good-quality mass spectra. CE-MS analysis of a reconstituted formulation of the biopharmaceutical recombinant human growth hormone (rhGH) which was stored for a prolonged time, revealed one degradation product which was provisionally identified as desamido rhGH. Based on the MS responses the amount of degradation was estimated to be ca. 25%.
ISSN:0003-2654
1364-5528
DOI:10.1039/b607178c