, Volume 34, Issue 1–2, pp 47–57 | Cite as

The effect of glucose and glutamine on the intracellular nucleotide pool and oxygen uptake rate of a murine hybridoma

  • N. Barnabé
  • M. ButlerEmail author


The effects of media concentrations of glucose andglutamine on the intracellular nucleotide pools andoxygen uptake rates of a murine antibody-secretinghybridoma cell line were investigated. Cells takenfrom mid-exponential phase of growth were incubated inmedium containing varying concentrations of glucose(0–25 mM) and glutamine (0–9 mM). The intracellularconcentrations of ATP, GTP, UTP and CTP, and theadenylate energy charge increased concomitantly withthe medium glucose concentration. The total adenylatenucleotide concentration did not change over a glucose concentration range of 1–25 mM but therelative levels of AMP, ADP and ATP changed as theenergy charge increased from 0.36 to 0.96. Themaximum oxygen uptake rate (OUR) was obtained in thepresence of 0.1–1 mM glucose. However at glucoseconcentrations >1 mM the OUR decreased suggestinga lower level of aerobic metabolism as a result of theCrabtree effect.A low concentration of glutamine (0.5 mM) caused asignificant increase (45–128%) in the ATP, GTP,CTP, UTP, UDP-GNac, and NAD pools and a doubling ofthe OUR compared to glutamine-free cultures. Theminimal concentration of glutamine also caused anincrease in the total adenylate pool indicating thatthe amino acid may stimulate thede novosynthesis of nucleotides. However, all nucleotidepools and the OUR remained unchanged within the rangeof 0.5–9 mM glutamine.Glucose was shown to be the major substrate forenergy metabolism. It was estimated that in thepresence of high concentrations of glucose (10–25 mM),glutamine provided the energy for the maintenance ofup to 28% of the intracellular ATP pool, whereas theremainder was provided by glucose metabolism.

glucose glutamine hybridoma nucleotides oxygen 


Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.


  1. Ardawi MSM and Newsholme EA (1983) Glutamine metabolism in lymphocytes of the rat. Biochem J 212: 835–842.Google Scholar
  2. Atkinson DE (1977) Cellular Energy Metabolism and its Regulation. Academic Press, New York, pp. 85–107 and pp. 200–224.Google Scholar
  3. Barnabé N and Butler M (1994) Effect of temperature on nucleotide pools and monoclonal antibody production in a mouse hybridoma. Biotechnol Bioeng 44: 1235–1245.Google Scholar
  4. Barnabé N and Butler M (1998) The relationship between intracellular UDP-N-acetyl hexosamine nucleotide pool and monoclonal antibody production in a mouse hybridoma. J Biotechnol 60: 67–80.Google Scholar
  5. Butler M and Jenkins HA (1989) Nutritional aspects of the growth of animal cells in culture. J Biotechnol 12: 97–110.Google Scholar
  6. Christie A and Butler M (1999) The adaptation of BHK cells to a non-ammoniagenic glutamate-based culture medium. Biotechnol Bioeng 64: 298–309.Google Scholar
  7. De Korte D, Haverkort WA, de Boer M, van Gennip AH and Roos D (1987) Imbalance in the nucleotide pools of myeloid leukemia cells and HL-60 cells: correlation with cell cycle phase, proliferation, differentiation and transformation. Cancer Res 47: 1841–1847.Google Scholar
  8. Dell'Antone P (1994) Metabolic pathways in Ehrlich ascites tumor cells recovering from a low bioenergetic status. FEBS Lett 350: 183–186.Google Scholar
  9. Engstrom W and Zetterberg A (1984) The relationship between purines, pyrimidines, nucleotides, and glutamine for fibroblast cell proliferation. J Biol Chem 256: 7812–7819.Google Scholar
  10. Fitzpatrick L, Jenkins HA and Butler M (1993) Glucose and glutamine metabolism of a murine B-lymphocyte hybridoma grown in batch culture. Appl Biochem Biotechnol 43: 93–116.Google Scholar
  11. Frenes SE, Furukawa RD, Li RK, Tumiati LC, Wersel RD and Mickle DA (1992) In vitro assessment of the effects of glucose added to the University of Wisconsin solution on myocyte preservation. Circulation 86 (5 Suppl.): II289-II294.Google Scholar
  12. Glacken MW (1988) Catabolic control of mammalian cell culture. Bio/Technology 6: 1041–1050.Google Scholar
  13. Grammatikos SI, Valley U, Nimtz M, Conradt HS and Wagner R (1998) Intracellular UDP-N-acetylhexosamine pool affects N-glycan complexity: A mechanism of ammonium action on protein glycosylation. Biotechnol Prog 14: 410–419.Google Scholar
  14. Guppy M, Abas L, Neylon C, Whisson ME, Whitham S, Pethwick DW and Niu X (1997) Fuel choices by human platelets in human plasma. Eur J Biochem 244: 161–167.Google Scholar
  15. Henderson JF and Paterson ARP (1973) Nucleotide Metabolism: An Introduction. Academic Press, New York, pp. 28–56.Google Scholar
  16. Jan DCH, Petch DA, Huzel N and Butler M (1997) The effect of dissolved oxygen on the metabolic profile of a murine hybridoma grown in serum-free medium in continuous culture. Biotechnol Bioeng 54: 153–164.Google Scholar
  17. Lanks KW and Li PW (1988) End products of glucose and glutamine metabolism by cultured cell lines. J Cell Physiol 135: 151–155.Google Scholar
  18. Lehninger AL (1982) Principles of Biochemistry. Worth Publishers, Inc., New York, 1011 p.Google Scholar
  19. Lomax CA and Henderson JF (1973) Adenosine formation and metabolism during adenosine triphosphate catabolism in Ehrlich ascites. Cancer Res. 33: 2825–2829.Google Scholar
  20. Lund P (1985) L-glutamine and L-glutamate. In: HU Bergmeyer (ed), Methods of Enzymatic Analysis, 3rd ed., VCH Verlagsgesellschaft, Weinheim, pp. 357–363.Google Scholar
  21. Lundin A, Hasenson M, Persson J and Pousette A (1986) Estimation of biomass in growing cell lines by adenosine triphosphate assay. Met. Enz. 133: 27–42.Google Scholar
  22. McComb RB and Yushok WD (1964) Metabolism of ascites tumor cells IV. Enzymatic reactions involved in adenosine triphosphate degradation induced by 2-deoxyglucose. Cancer Res 24: 198–203.Google Scholar
  23. McKeehan WL (1982) Glycolysis, glutaminolysis and cell proliferation. Cell Biol Int Rep 6: 635–650.Google Scholar
  24. Medina MA and Nunez de Castro I (1990) Glutaminolysis and glycolysis interactions in proliferant cells. Int J Biochem 22: 681–683.Google Scholar
  25. Meijer JJ and van Dijken JP (1995) Effects of glucose supply on myeloma growth and metabolism in chemostat culture. J Cell Physiol 162: 191–198.Google Scholar
  26. Miller WM, Wilke CR and Blanch HW (1989a) Transient responses of hybridoma cells to nutrient additions on continuous culture: I. Glucose pulse and step changes. Biotechnol Bioeng 33: 447–486.Google Scholar
  27. Miller WM, Wilke CR and Blanch HW (1989b) The transient responses of hybridoma cells to nutrient additions on continuous culture: II. Glutamine pulse and step changes. Biotechnol Bioeng 33: 487–499.Google Scholar
  28. Neermann J and Wagner R (1996) Comparative analysis of glucose and glutamine metabolism in transformed mammalian cell lines, insect and primary liver cells. J Cell Physiol 166: 152–169.Google Scholar
  29. Petch D and Butler M (1994) A profile of energy metabolism in a murine hybridoma: glucose and glutamine utilization. J Cell Physiol 161: 71–76.Google Scholar
  30. Reitzer LJ, Wice BM and Kennell D (1980) The pentose cycle: control and essential function in HeLa cell nucleic acid synthesis. J Biol Chem 255: 5616–5626.Google Scholar
  31. Ronca-Testoni S and Ronca G (1974) Muscle 51-adenylic acid aminohydrolase. Kinetic properties of rat muscle enzyme treated with pyridoxal 50-phosphate. J Biol Chem 249: 7723–7728.Google Scholar
  32. Ryll T and Wagner R (1992) Intracellular ribonucleotide pools as a tool for monitoring the physiological state of in vitro cultivated mammalian cells during production processes. Biotechnol Bioeng 40: 934–946.Google Scholar
  33. Ryll T, Valley U and Wagner R (1994) Biochemistry of growth inhibition by ammonium ions in mammalian cells. Biotechnol Bioeng 44: 184–193.Google Scholar
  34. Sri-Pathmanathan RM, Braddock P and Brindle KM (1990)31PNMR studies of glucose and glutamine metabolism in cultured mammalian cells. Biochim Biophys Acta 1051: 131–137.Google Scholar
  35. Wohlpart D, Kirman D and Gainer J (1990) Effects of cell density and glucose and glutamine levels on the respiration rates of hybridoma cells. Biotechnol Bioeng 36: 630–635.Google Scholar
  36. Zielke HR, Ozand PT, Tildon JT, Sevdalian DA and Cornblath M (1976) Growth of human diploid fibroblasts in the absence of glucose utilization. Proc Natl Acad Sci USA 73: 4110–4114.Google Scholar

Copyright information

© Kluwer Academic Publishers 2000

Authors and Affiliations

  1. 1.Department of MicrobiologyUniversity of Manitoba, WinnipegManitobaCanada
  2. 2.Department of MicrobiologyUniversity of Manitoba, WinnipegManitobaCanada

Personalised recommendations