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Analysis of cytokinin nucleotides by capillary zone electrophoresis with diode array and mass spectrometric detection in a recombinant enzyme in vitro reaction
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SYSNO ASEP 0385991 Document Type J - Journal Article R&D Document Type Journal Article Subsidiary J Článek ve WOS Title Analysis of cytokinin nucleotides by capillary zone electrophoresis with diode array and mass spectrometric detection in a recombinant enzyme in vitro reaction Author(s) Béres, Tibor (UEB-Q) RID
Gemrotová, Markéta (UEB-Q)
Tarkowski, P. (CZ)
Ganzera, M. (AT)
Maier, V. (CZ)
Fridecký, D. (CZ)
Dessoy, M. A. (DE)
Wessjohann, L. A. (DE)
Spíchal, Lukáš (UEB-Q) RID, ORCID
Strnad, Miroslav (UEB-Q) RID, ORCID
Doležal, Karel (UEB-Q) RID, ORCIDSource Title Analytica Chimica Acta. - : Elsevier - ISSN 0003-2670
Roč. 751, Feb (2012), s. 176-181Number of pages 6 s. Language eng - English Country NL - Netherlands Keywords Cytokinin nucleotides ; Capillary electrophoresis ; Isopentenyltransferase Subject RIV ED - Physiology R&D Projects LC06034 GA MŠMT - Ministry of Education, Youth and Sports (MEYS) CEZ AV0Z50380511 - UEB-Q (2005-2011) UT WOS 000311013100021 DOI 10.1016/j.aca.2012.08.049 Annotation A capillary zone electrophoresis (CZE) method for separation of adenosine and N-6-isopentenyladenosine (cytokinin) nucleotides was developed, optimized and validated. Aqueous solutions of several amino acids were evaluated as the background electrolyte constituents. Separation of six nucleotides in less than 20 min with high theoretical plate number (up to 400 000 for isopentenyladenosine triphosphate) was achieved using a 100 mM sarcosine/ammonia buffer at pH 10.0. The detection limits of the CZE-UV method are in the low micromolar range (0.69-1.27 mu mol L-1). Good repeatability of migration times (within 1.3%), peak areas (within 1.8%) and linearity (R-2 >0.999) was achieved over the concentration range 5-1000 mu mol L-1. The method was used to assay the activity of the recombinant Arabidopsis thaliana isopentenyltransferase 1 (AtIPT1). Baseline separation of isopentenylated nucleotides by CE-ESI-MS using a volatile buffer (30 mM ammonium formate; pH 10.0) was accomplished. The identities of the reaction products - isopentenyladenosine di- and triphosphate were confirmed by HPLC-QqTOE-MS. Dephosphorylation of ATP was observed as a parallel reaction. Workplace Institute of Experimental Botany Contact David Klier, knihovna@ueb.cas.cz, Tel.: 220 390 469 Year of Publishing 2013
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