Advertisement

Cytotechnology

, Volume 14, Issue 2, pp 123–128 | Cite as

A simple HPLC technique for accurate monitoring of mammalian cell metabolism

  • T. Stoll
  • P. Pugeaud
  • U. von Stockar
  • I. W. Marison
Technical Report

Abstract

We have developed a simple and accurate isocratic HPLC method, without any prederivatisation, for the determination of glucose, lactate, glutamine, glutamate, pyrrolidone carboxylic acid and alanine in samples from mammalian cell cultures. The method has been successfully validated with enzyme analysis for each of the compounds. Quantification of pyrrolidone carboxylic acid makes the correction for glutamine decrease due to chemical decomposition very simple and accurate, and avoids some possibly erroneous calculations.

Key words

HPLC mammalian cells metabolism pyrrolidone carboxylic acid 

Preview

Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.

References

  1. Devore JL (1982) Probability and Statistics for Engineering and the Sciences. Brooks/Cole Publishing Co, Wadsworth Inc., Monterey CA.Google Scholar
  2. Favre E, Pugeaud P, Péringer P (1990) Automated HPLC monitoring of glucose, glutamine, lactate and alanine on suspended mammalian cell reactors. Biotechnol. Tech. 4: 5, 315–320.Google Scholar
  3. Gilbert JB, Price VE, Greenstein JP (1949) Effect of anions on the non-enzymatic desamidation of glutamine. J. Biol. Chem. 180: 209–218.Google Scholar
  4. Hassel T, Gleave S, Butler M (1991) Growth inhibition in animal cell culture: The effect of lactate and ammonia. Appl. Biochem. Biotechnol. 30: 29–41.Google Scholar
  5. Ozturk SS, Palsson BO (1990) Chemical decomposition of glutamine in cell culture media: Effect of media type, pH, and serum concentration. Biotechnol. Prog. 6: 121–128.Google Scholar
  6. Tritsch GL and Moore GE (1962) Spontaneous decomposition of glutamine in cell culture media. Exp. Cell Res. 28: 360–364.Google Scholar
  7. Weigang F, Reiter M, Jungbauer A, Katinger H (1989) Highperformance liquid chromatographic determination of metabolic production for fermentation control of mammalian cell culture: Analysis of carbohydrates, organic acids and orthophosphate using refractive index and ultraviolet detectors. J. Chromatography 497: 59–68.Google Scholar
  8. Wein J and Götz IE (1973) Asparaginase and glutaminase activities in culture media containing dialyzed fetal calf serum. In Vitro 9: 3, 186–193.Google Scholar
  9. Williamson DH (1974) L-Alanin: Bestimmung mit Alanin-Dehydrogenase. In: HU Bergmeyer (ed.) Methoden der Enzymatischen Analyse. Band I. (pp. 1724–1727) 3rd edition. Verlag Chemie, Weinheim.Google Scholar

Copyright information

© Kluwer Academic Publishers 1994

Authors and Affiliations

  • T. Stoll
    • 1
  • P. Pugeaud
    • 1
  • U. von Stockar
    • 1
  • I. W. Marison
    • 1
  1. 1.Institute of Chemical EngineeringSwiss Federal Institute of Technology (EPFL)LausanneSwitzerland

Personalised recommendations