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Ion-pair reversed phase HPLC determination of nucleotides, nucleosides and nucleobases — Application to nucleotide metabolism in hepatocytes

  • Late Papers from the 17th International Symposium on Chromatography, Vienna, Austria, September 25–30, 1988
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Three groups of metabolites were analyzed in extracts of rat hepatocytes by an HPLC method: (i) nucleotides (ATP, ADP, AMP, GTP, GDP, UTP, UDP, IMP, UMP), (ii) nucleosides and nucleobases (adenosine, adenine, guanosine, inosine, xanthine, hypoxanthine, uridine) and (iii) inhibitors of xanthine oxidoreductase (oxypurinol, allopurinol). Perchloric acid extracts were neutralized with K2CO3/triethanolamine and analyzed at 254 nm by reversed-phase ion-pair high-performance liquid chromatography. The nucleotides and their derivatives were separated with a gradient elution using 10 mM NH4H2PO4/2 mM t-butylammonium-phosphate and acetonitrile. Recovery values were estimated for the extraction procedure used. The method was applied to the investigation of nucleotide metabolism of hepatocytes of starved rats at anoxia and reoxygenation.

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Werner, A., Schneider, W., Siems, W. et al. Ion-pair reversed phase HPLC determination of nucleotides, nucleosides and nucleobases — Application to nucleotide metabolism in hepatocytes. Chromatographia 27, 639–643 (1989).

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