Quantitation of intracellular purine intermediates in different Corynebacteria using electrospray LC-MS/MS

Susanne Peifer, Konstantin Schneider, Gudrun Nuerenberg, Dietrich A. Volmer, Elmar Heinzle

Research output: Contribution to journalJournal articleResearchpeer-review


Intermediates of the purine biosynthesis pathway play key roles in cellular metabolism including nucleic acid synthesis and signal mediation. In addition, they are also of major interest to the biotechnological industry as several intermediates either possess flavor-enhancing characteristics or are applied in medical therapy. In this study, we have developed an analytical method for quantitation of 12 intermediates from the purine biosynthesis pathway including important nucleotides and their corresponding nucleosides and nucleobases. The approach comprised a single-step acidic extraction/quenching procedure, followed by quantitative electrospray LC-MS/MS analysis. The assay was validated in terms of accuracy, precision, reproducibility, and applicability for complex biological matrices. The method was subsequently applied for determination of free intracellular pool sizes of purine biosynthetic pathway intermediates in the two Gram-positive bacteria Corynebacterium glutamicum and Corynebacterium ammoniagenes. Importantly, no ion pair reagents were applied in this approach as usually required for liquid chromatography analysis of large classes of diverse metabolites.
Original languageEnglish
JournalAnalytical and Bioanalytical Chemistry
Issue number8
Pages (from-to)2295-2305
Number of pages11
Publication statusPublished - 2012
Externally publishedYes


  • Biological samples
  • HPLC
  • Mass spectrometry
  • Nucleic acid sampling
  • Quantitative extraction
  • Bioanalytical methods


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