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The effect of sample salt additives on capillary electrophoresis analysis of intact proteins using surface modified capillaries.
Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry, Analytical Chemistry. Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry.
Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry, Analytical Chemistry. Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry.
Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry, Analytical Chemistry. Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry.
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
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2009 (English)In: Journal of Chromatography A, ISSN 0021-9673, E-ISSN 1873-3778, Vol. 1216, no 17, 3613-3620 p.Article in journal (Refereed) Published
Abstract [en]

The effect of adding alkali salts to protein samples for capillary electrophoretic (CE) analysis of intact proteins was studied. A high degree of peak stacking, even for large proteins, was found to occur when alkali salts were added to the sample. The addition of salt to the protein sample promotes a strong improvement in the peak efficiency of individual proteins giving up to 2.1 x 10(6) apparent plates/m. The concentration of salt required in the sample to reach optimal peak efficiency show dependency on both the molecular weight and molar concentration of the protein. However, adding salt will, at a sufficiently high concentration, cause a mixture of proteins to co-migrate to one very sharp peak. The observed sample stacking effect was obtained with a number of different surface modified silica capillaries indicating a general phenomenon and not surface coating specific.

Place, publisher, year, edition, pages
2009. Vol. 1216, no 17, 3613-3620 p.
Keyword [en]
Capillary electrophoresis, alkali salt, intact protein analysis, coated capillary, stacking effect
National Category
Chemical Sciences
Research subject
Analytical Chemistry
Identifiers
URN: urn:nbn:se:uu:diva-100628DOI: 10.1016/j.chroma.2008.12.037ISI: 000265467200004PubMedID: 19150070OAI: oai:DiVA.org:uu-100628DiVA: diva2:210704
Available from: 2009-04-03 Created: 2009-04-03 Last updated: 2011-05-13Bibliographically approved
In thesis
1. Development of Capillary Electrophoresis Methods Coupled to Mass Spectrometry for Biomedical and Pharmaceutical Analysis
Open this publication in new window or tab >>Development of Capillary Electrophoresis Methods Coupled to Mass Spectrometry for Biomedical and Pharmaceutical Analysis
2011 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

The analysis of large intact proteins and complex biological samples containing drug molecules is a common complicated task for many scientists. However, due to the importance of these molecules, there is a growing interest in pharmaceutical and medicinal research to develop rapid, highly sensitive and efficient analytical techniques. The advantages of capillary electrophoresis (CE) in combination with mass spectrometry (MS) provide a powerful analytical tool. However, further improvement and development of these techniques are required to extend their utility and to meet the challenges of selected analytes. Thus, the scope of this thesis deals with the development of novel analytical methods to achieve efficient and high performance analysis of peptides, intact proteins, digests of complex samples and basic pharmaceutical drug compounds in biological matrices.

Implementation of CE for routine analysis of proteins and complex samples is constrained by the partial adsorption to the capillary wall. Consequently, the use of surface modified capillaries is required to control the surface properties and prevent analyte adsorption. In this thesis, analyte adsorption was successfully prevented using tailored covalent cationic (M7C4I) and electrostatic cationic (PVPy-Me) coatings. Rapid and efficient separations of peptides, proteins and digests of complex samples such as cerebrospinal fluids were obtained with these coatings. The M7C4I coating showed a distinct ability to handle large intact proteins with a molecular size of over 0.5 MDa. The highest peak efficiencies and surprisingly high peak stacking effects were obtained by adding salts to the protein samples. The effect of salt additives on peak efficiencies of intact proteins was further demonstrated and compared using different surface modified capillaries. Additionally, rapid CE-ESI-MS quantification of pharmaceutical drug molecules in human plasma was performed after a SCX-SPE sample preparation method using the M7C4I coating. In conclusion, the results presented in this thesis show the strong potential of CE in combination with MS using electrospray ionization (ESI) for the analysis of peptides and large intact proteins and the applicability for clinical monitoring of the levels of pharmaceutical drug molecules in human plasma with high sensitivity and efficiency.

Place, publisher, year, edition, pages
Uppsala: Acta Universitatis Upsaliensis, 2011. 64 p.
Series
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Science and Technology, ISSN 1651-6214 ; 804
Keyword
Capillary Electrophoresis, Capillary Surface Modificaions, Electrospray Ionization, Mass Spectrometry, Peptides, Intact Proteins, Basic Pharmaceutical Drug Molecules and Complex Biological Samples
National Category
Other Basic Medicine
Research subject
Analytical Chemistry
Identifiers
urn:nbn:se:uu:diva-143814 (URN)978-91-554-7996-1 (ISBN)
Public defence
2011-03-11, C8:301, BMC, Husargatan 3, Uppsala, 13:15 (English)
Opponent
Supervisors
Note
Felaktigt tryckt som Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Science and Technology 734Available from: 2011-02-18 Created: 2011-01-25 Last updated: 2011-04-04Bibliographically approved

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Wetterhall, MagnusBergquist, Jonas

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