Determination of δ-[ L -α-aminoadipyl]- L -cysteinyl- D -valine in cell extracts of Penicillium chrysogenum using ion pair-RP-UPLC-MS/MS

Reza Maleki Seifar*, Amit T. Deshmukh, Joseph J. Heijnen, Walter M. Van Gulik

*Corresponding author for this work

    Research output: Contribution to journalArticleScientificpeer-review

    4 Citations (Scopus)

    Abstract

    δ-[L-α-Aminoadipyl]-L-cysteinyl-D-valine (ACV) is a key intermediate in the biosynthesis pathway of penicillins and cephalosporins. Therefore, the accurate quantification of ACV is relevant, e.g. for kinetic studies on the production of these β-lactam antibiotics. However, accurate quantification of ACV is a challenge, because it is an active thiol compound which, upon exposure to air, can easily react with other thiol compounds to form oxidized disulfides. We have found that, during exposure to air, the oxidation of ACV occurs both in aqueous standard solutions as well as in biological samples. Qualitative and quantitative determinations of ACV and the oxidized dimer bis-δ-[L-α-aminoadipyl]-L-cysteinyl-D-valine have been carried out using ion pair reversed-phase ultra high-performance liquid chromatography, hyphenated with tandem mass spectrometry (IP-RP-UPLC-MS/MS) as the analytical platform. We show that by application of tris(2-carboxy-ethyl)phosphine hydrochloride (TCEP) as the reducing reagent, the total amount of ACV can be determined, while using maleimide as derivatizing reagent enables to quantify the free reduced form only.

    Original languageEnglish
    Pages (from-to)225-230
    Number of pages6
    JournalJournal of Separation Science
    Volume35
    Issue number2
    DOIs
    Publication statusPublished - Jan 2012

    Keywords

    • Cell extract
    • Maleimide
    • Penicillium chrysogenum
    • TCEP
    • Thiols

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