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Derivatisation for liquid chromatography/electrospray mass spectrometry: synthesis of pyridinium compounds and their amine and carboxylic acid derivatives

Authors

  • Samantha J. Barry,

    1. Chemical Development, GlaxoSmithKline Research and Development Ltd., New Frontiers Science Park North, Third Avenue, Harlow CM19 5AW, UK
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  • Richard M. Carr,

    1. Department of Isotope Chemistry, Discovery Research, GlaxoSmithKline Research and Development Ltd., Gunnels Wood Road, Stevenage, SG1 2NY, UK
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  • Stephen J. Lane,

    1. Analytical Sciences, Discovery Research, GlaxoSmithKline Research and Development Ltd., Gunnels Wood Road, Stevenage SG1 2NY, UK
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  • William J. Leavens,

    Corresponding author
    1. Analytical Sciences, Discovery Research, GlaxoSmithKline Research and Development Ltd., Gunnels Wood Road, Stevenage SG1 2NY, UK
    • Analytical Sciences, Discovery Research, GlaxoSmithKline Research and Development Ltd., Gunnels Wood Road, Stevenage SG1 2NY, UK.
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  • Soraya Monté,

    1. Chemical Development, GlaxoSmithKline Research and Development Ltd., New Frontiers Science Park North, Third Avenue, Harlow CM19 5AW, UK
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  • Ian Waterhouse

    1. Department of Isotope Chemistry, Discovery Research, GlaxoSmithKline Research and Development Ltd., Gunnels Wood Road, Stevenage, SG1 2NY, UK
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Abstract

A simple method has been developed for the pre-column derivatisation of low molecular weight primary and secondary amines and carboxylic acids using quaternary nitrogen compounds to enhance their detection by liquid chromatography/electrospray ionisation mass spectrometry (LC/ESI-MS). The synthesis of seven novel quaternary nitrogen reagents is described. The derivatives are designed to be relatively small molecules to avoid some of the steric hindrance problems that may be associated with larger derivatisation reagents. The compounds have amine and carboxylic acid functional groups with which to derivatise carboxylic acids and amines, respectively. Two of the compounds contain a bromine atom in order to assess the advantages of a bromine isotope pattern in the mass spectra. This acts as a simple marker for derivatisation and enables data processing by cluster analysis.

Activation of the carboxylic acid group was achieved by the use of either 1-chloro-4-methylpyridinium iodide (CMPI) or the more reactive 1-fluoro-4-methylpyridinium p-toluenesulphonate (FMP).1 Using both of these active reagents, the degree of nucleophilic substitution was investigated for the derivatisation of a variety of small molecules. Whilst giving some increase in the ESI-MS response for the derivatised compounds, the FMP itself acted as a derivatising reagent in a competing reaction. In the light of this finding, FMP was reacted with the test compounds separately and gave positive results as a derivatising reagent. Detection of the ‘pre-charged’ derivatives of amines and carboxylic acids by LC/ESI-MS was investigated with respect to their ESI response and chromatography. Copyright © 2003 John Wiley & Sons, Ltd.

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