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Tandem mass spectrometry identification and LC-MS quantification of intact cytokinin nucleotides in K-562 human leukemia cells
T. Béres, M. Zatloukal, J. Voller, P. Niemann, MC. Gahsche, P. Tarkowski, O. Novák, J. Hanuš, M. Strnad, K. Doležal,
Jazyk angličtina Země Německo
Typ dokumentu časopisecké články, práce podpořená grantem
- MeSH
- chromatografie kapalinová MeSH
- cytokininy analýza chemie MeSH
- hmotnostní spektrometrie MeSH
- lidé MeSH
- molekulární struktura MeSH
- nádorové buněčné linie MeSH
- nukleotidy analýza chemie MeSH
- tandemová hmotnostní spektrometrie MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
We describe here a new reversed-phase high-performance liquid chromatography with mass spectrometry detection method for quantifying intact cytokinin nucleotides in human K-562 leukemia cells. Tandem mass spectrometry was used to identify the intracellular metabolites (cytokinin monophosphorylated, diphosphorylated, and triphosphorylated nucleotides) in riboside-treated cells. For the protein precipitation and sample preparation, a trichloroacetic acid extraction method is used. Samples are then back-extracted with diethyl ether, lyophilized, reconstituted, and injected into the LC system. Analytes were quantified in negative selected ion monitoring mode using a single quadrupole mass spectrometer. The method was validated in terms of retention time stabilities, limits of detection, linearity, recovery, and analytical accuracy. The developed method was linear in the range of 1-1,000 pmol for all studied compounds. The limits of detection for the analytes vary from 0.2 to 0.6 pmol.
Citace poskytuje Crossref.org
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- $a Béreš, Tibor $u Laboratory of Growth Regulators, IEB AS CR and Palacký University, Šlechtitelů 11, 78371 Olomouc, Czech Republic. $7 xx0119829
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- $a We describe here a new reversed-phase high-performance liquid chromatography with mass spectrometry detection method for quantifying intact cytokinin nucleotides in human K-562 leukemia cells. Tandem mass spectrometry was used to identify the intracellular metabolites (cytokinin monophosphorylated, diphosphorylated, and triphosphorylated nucleotides) in riboside-treated cells. For the protein precipitation and sample preparation, a trichloroacetic acid extraction method is used. Samples are then back-extracted with diethyl ether, lyophilized, reconstituted, and injected into the LC system. Analytes were quantified in negative selected ion monitoring mode using a single quadrupole mass spectrometer. The method was validated in terms of retention time stabilities, limits of detection, linearity, recovery, and analytical accuracy. The developed method was linear in the range of 1-1,000 pmol for all studied compounds. The limits of detection for the analytes vary from 0.2 to 0.6 pmol.
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