Reversed-phase high-performance liquid chromatography of purine compounds for investigation of biomedical problems: application to different tissues and body fluids

Grune, Tilman; Siems, Werner

doi:10.1016/0378-4347(93)80025-y

Cited by 26 publications

(6 citation statements)

References 74 publications

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“…After 2-deoxyglucose/ azide treatment, we did not see a corresponding increase in any of the nucleotides (such as AMP) that might possibly represent breakdown or interconversion products of ATP (unpublished data). This is consistent with a previous report that a decrease in cellular ATP stimulated by anoxia correlates with a rise at time points similar to those observed here in the metabolic breakdown products of ATP, such as adenosine, that are not retained on our FPLC columns (Grune and Siems, 1993).…”

Section: Total Nucleotide Levels In Atp-and Gtp-depleted Cellssupporting

confidence: 93%

The mechanism of inhibition of Ran-dependent nuclear transport by cellular ATP depletion

Schwoebel

Moore

2002

The Journal of Cell Biology

121

120

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Rran-dependent nuclear transport requires a nuclear pool of RanGTP both for the assembly of export complexes and the disassembly of import complexes. Accordingly, in order for these processes to proceed, Ran-dependent nuclear import and export assays in vitro require the addition of GTP to produce RanGTP. Notably, no ATP requirement can be detected for these transport processes in vitro. But in vivo, when cells are depleted of ATP by the addition of sodium azide and 2-deoxyglucose to block ATP production by oxidative phosphorylation and glycolysis, respectively, Ran-dependent nuclear import and export are rapidly inhibited. This raised the question of whether there is an ATP requirement for these nuclear transport pathways in an intact cell that has remained undetected in vitro. Here we report that the free (but not total) GTP concentration rapidly drops to an undetectable level upon ATP depletion as does the availability of RanGTP. Our conclusion is that the inhibition of Ran-dependent nuclear transport observed upon ATP depletion in vivo results from a shortage of RanGTP rather than the inhibition of some ATP-dependent process.

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Section: Total Nucleotide Levels In Atp-and Gtp-depleted Cellssupporting

confidence: 93%

The mechanism of inhibition of Ran-dependent nuclear transport by cellular ATP depletion

Schwoebel

Moore

2002

The Journal of Cell Biology

121

120

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“…In whole blood, precipitate volumes are large, and the 20% adjustment of concentration to correct for this volume change may have been an overcorrection for more apolar metabolites, such as uric acid, adenine and hypoxanthine, resulting in lower recoveries. Recovery levels reported by others for methods using perchloric acid for the extraction of nucleotides from cells or tissues range between 75 and 120.5% [27][28][29][30]. The recoveries did not differ substantially whether the nucleotides were spiked at high or low concentrations.…”

Section: Discussionmentioning

confidence: 80%

Simultaneous determination of adenosine triphosphate and its metabolites in human whole blood by RP-HPLC and UV-detection

Coolen

Arts

Swennen

et al. 2008

Journal of Chromatography B

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“…Fürst and Hallström, 1992) could face difficulties in proper peak resolution, the separation of nucleotides by ion-pairing HPLC could be achieved nearly in all conditions. Moreover, care should be taken to avoid too tight binding of the nucleotides in the column rather than make efforts to achieve properly resolved peaks for the species: it has been shown that dozens of nucleotide metabolites could be resolved in a single run (see Grune and Siems, 1993 for review).…”

Section: Previous Studies and Uplcmentioning

confidence: 99%

“…Brown et al, 1980;Grune and Siems, 1993) as described in Materials and Methods. The risk factors for the procedure are (1) instability of PCr as well as Cr at (highly) acidic pH values (see, e.g.…”

Section: Sample Preparationmentioning

confidence: 99%

“…In the ion-pair HPLC method, the separation of (negatively) charged compounds is expectedly achieved due to (1) the modification of the column's surface by reversible binding of large hydrophobic cations, e.g., TBA; and (2) the binding of the complex of the cation with oppositely charged species formed in the mobile phase to the hydrophobic stationary phase (see, e.g., Grune and Siems, 1993). Nevertheless, for nucleotides as well as Cr and PCr only the first mechanism seems to hold as it was generally accepted years ago Alberty, 1956a, 1956b) that TBA cation does not associate with phosphate-containing ligands in water solutions.…”

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confidence: 99%

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Ultra performance liquid chromatography analysis of adenine nucleotides and creatine derivatives for kinetic studies

Sikk¹,

Käämbre²,

Vija³

et al. 2009

Proc. Estonian Acad. Sci.

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A rapid method for simultaneous quantification of compounds participating in energy metabolism of cardiac muscle cells (creatine (Cr), phosphocreatine (PCr), ADP, and ATP) is described where a conventional ion-pair reversed phase HPLC separation has been improved by introducing the method based on the recently developed ultra performance liquid chromatography (UPLC) technique. In the 0.005-1 mM concentration range, the calibration curves for Cr, ADP, and ATP as pure standard compounds were fitted by the polynomic relationship y = y 0 + ax -bx 2 at a high confidence level (R 2 > 0.999 in all cases) and that for PCr by a simple linear relationship with R 2 > 0.998. The method was applied for the study of the kinetics of PCr production by permeabilized cardimyocytes due to the cellular oxydative phosphorylation reactions. The determined steady-state levels of ADP and ATP as well as the rate of PCr production in different conditions can be used for the verification of the results of mathematical modelling of cardiomyocyte functioning.

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Reversed-phase high-performance liquid chromatography of purine compounds for investigation of biomedical problems: application to different tissues and body fluids

Cited by 26 publications

References 74 publications

The mechanism of inhibition of Ran-dependent nuclear transport by cellular ATP depletion

The mechanism of inhibition of Ran-dependent nuclear transport by cellular ATP depletion

Simultaneous determination of adenosine triphosphate and its metabolites in human whole blood by RP-HPLC and UV-detection

Ultra performance liquid chromatography analysis of adenine nucleotides and creatine derivatives for kinetic studies

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