Simultaneous quantification of free nucleotides in complex biological samples using ion pair reversed phase liquid chromatography isotope dilution tandem mass spectrometry

A new sensitive and accurate analytical method has been developed for quantification of intracellular nucleotides in complex biological samples from cultured cells of different microorganisms such as Saccharomyces cerevisiae, Escherichia coli, and Penicillium chrysogenum. This method is based on ion...

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Bibliographic Details
Published in:Analytical biochemistry 2009-05, Vol.388 (2), p.213-219
Main Authors: Seifar, Reza M., Ras, Cor, van Dam, Jan C., van Gulik, Walter M., Heijnen, Joseph J., van Winden, Wouter A.
Format: Article
Language:English
Subjects:
Chromatography, Liquid - methods
Escherichia coli
Escherichia coli - genetics
Intracellular nucleotides
Ion pair reversed phase liquid chromatography
Isotope dilution mass spectrometry
Isotopes - analysis
Nucleotides - analysis
Nucleotides - chemistry
Penicillium chrysogenum
Penicillium chrysogenum - genetics
Saccharomyces cerevisiae
Saccharomyces cerevisiae - genetics
Spectrometry, Mass, Electrospray Ionization
Tandem Mass Spectrometry - methods
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Summary:A new sensitive and accurate analytical method has been developed for quantification of intracellular nucleotides in complex biological samples from cultured cells of different microorganisms such as Saccharomyces cerevisiae, Escherichia coli, and Penicillium chrysogenum. This method is based on ion pair reversed phase liquid chromatography electrospray ionization isotope dilution tandem mass spectrometry (IP–LC–ESI–ID–MS/MS. A good separation and low detection limits were observed for these compounds using dibutylamine as volatile ion pair reagent in the mobile phase of the LC. Uniformly 13C-labeled isotopes of nucleotides were used as internal standards for both extraction and quantification of intracellular nucleotides. The method was validated by determining the linearity, sensitivity, and repeatability.
ISSN:0003-2697
1096-0309
DOI:10.1016/j.ab.2009.02.025