Abstract
Although glutamine is used as a major substrate for the growth of mammalian cells in culture, it suffers from some disadvantages. Glutamine is deaminated through storage or by cellular metabolism, leading to the formation of ammonia which can result in growth inhibition. Non-ammoniagenic alternatives to glutamine have been investigated in an attempt to develop strategies for obtaining improved cell yields for ammonia sensitive cell lines.
Glutamate is a suitable substitute for glutamine in some culture systems. A period of adaptation to glutamate is required during which the activity of glutamine synthetase and the rate of transport of glutamate both increase. The cell yield increases when the ammonia accumulation is decreased following culture supplementation with glutamate rather than glutamine. However some cell lines fail to adapt to growth in glutamate and this may be due to a low efficiency transport system.
The glutamine-based dipeptides, ala-gln and gly-gln can substitute for glutamine in cultures of antibody-secreting hybridomas. The accumulation of ammonia in these cultures is less and cell yields in dipeptide-based media may be improved compared to glutamine-based controls. In murine hybridomas, a higher concentration of gly-gln is required to obtain comparable cell growth to ala-gln or gin-based cultures. This is attributed to a requirement for dipeptide hydrolysis catalyzed by an enzyme with higher affinity for ala-gln than gly-gln.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
Preview
Unable to display preview. Download preview PDF.
References
Ardawi MSM and Newsholme EA (1983) Glutamine metabolism in lymphocytes of the rat. Biochem. J. 212: 835–842.
Butler M, Imamura T, Thomas J and Thilly WG (1983) High yields from microcarrier cultures by medium perfusion. J. Cell Sci. 61: 351–363.
Butler M and Spier RE (1984) The effects of glutamine utilisation and ammonia production on the growth of BHK cells in microcarrier cultures. J. Biotech. 1: 187–196.
Butler M, Hassell T, Doyle C, Gleave S and Jennings P. (1991) The effect on metabolic by products on animal cells in culture. In: Spier RE, Griffiths JB and Meigner B (eds.) Production of biologicals from animal cells in Culture, ES ACT 10 (pp. 226–228) Butterworth-Heinemann, Oxford.
Christie A and Butler M (1994) Growth and metabolism of a murine hybridoma in cultures containing glutamine-based dipeptides. Focus 16: 9–13.
Doyle C and Butler M (1990) The effect of pH on the toxicity of ammonia to a murine hybridoma. J. Biotech. 15: 91–100.
Eagle H (1955) Nutrition needs of mammalian cells in tissue culture. Science 122: 501–504.
Eagle H, Oyama VI, Levy M, Horton CL and Fleischman R (1956) The growth response of mammalian cells in tissue culture to L-glutamine and L-glutamic acid. J. Biol. Chem. 218: 607–616.
Feng B, Shiber SK and Max SR (1990) Glutamine regulates glutamine synthetase expression in skeletal muscle cells in culture. J. Cell Physiol. 145: 376–380.
Glacken MW, Fleischaker, RJ and Sinskey, AJ (1986) Reduction of waste product excretion via nutrient control: possible strategies for maximising product and cell yields in cultures of mammalian cells. Biotech. Bioeng. 28: 1376–1389.
Griffiths JB (1973) The effects of adapting human diploid cells to grow in glutamic acid media on cell morphology, growth and metabolism. J. Cell Sci. 12: 617–629.
Hassell T and Butler M (1990) Adaptation to non-ammoniagenic medium and selective substrate feeding lead to enhanced yields in animal cell cultures. J. Cell Sci. 96: 501–508.
Hassell T, Gleave S and Butler M (1991) Growth inhibition in animal cell culture: the effect of lactate and ammonia. Appl. Biochem. Biotech. 30: 29–41.
Holmlund A-C, Chatzisavido N, Bell SL and Lindner-Olsson E (1992) Growth and metabolism of recombinant CHO cell-lines in serum-free medium containing derivatives of glutamine. In: Spier RE, Griffiths JB and MacDonald C (eds) Animal cell technology: developments, processes and products ES ACT 11 (pp. 176–179) Butterworth-Heinemann, Oxford.
Jenkins HA, Butler M and Dickson AJ (1992) Characterization of the importance of glutamine metabolism to hybridoma cell growth and productivity. J. Biotechnol. 23: 167–182.
Low SY, Rennie MJ, Taylor PM (1994) Sodium-dependent glutamate transport in cultured rat myotubes increases after glutamine deprivation. FASEB Journal 8: 127–131.
Ljunggren, J and Haggstrom L (1990) Glutamine Limited Fed-Batch Culture Reduces Ammonium Ion Production in Animal Cells. Biotechnol. Lett. 12: 705–710.
McDermott RH and Butler M (1993) Uptake of glutamate, not glutamine synthetase, regulates adaptation of mammalian cells to glutamine-free medium. J. Cell Sci. 104: 51–58.
Minamoto Y, Ogawa K, Abe H, Iochi Y and Mitsugi K (1991) Development of a serum-free and heat-sterilizable medium and continuous high-density culture. Cytotechnology 5, S35–51.
Reitzer LJ, Wice BM and Kennell D (1979) Evidence that glutamine not sugar is the major energy source for cultured HeLa cells. J. Biol. Chem. 254: 2669–2676.
Reuveny S, Velez D, Miller L and Macmillan JD (1986) Factors affecting cell growth and monoclonal antibody production in stirred reactors. J. Immunol. Methods 86: 53–59.
Roth E, Ollenschlager G, Hamilton G, Simmel A, Langer K, Fekyl W and Jakesz R (1988) Influence of two glutamine containing dipeptides on growth of mammalian cells In Vitro Cell. Develop. Biol. 24: 696.
Ryan WL and Cardin C (1966) Amino acids and ammonia of fetal calf serum during storage. Proc. Soc. Exp. Biol. Med. 123: 27–30.
Taya M, Mano T and Koybayashi, T (1986) Kinetic expression of human cell growth in a suspension culture system. J. Ferment. Technol. 64: 347–350.
Tritsch GL and Moore GE (1962) Spontaneous decomposition of glutamine in cell culture media. Expl. Cell Res. 28: 360–364.
Wice BM, Reitzer LJ and Kennell D (1981) The continuous growth of vertebrate cells in the absence of sugar. J. Biol. Chem. 256: 7812–7819.
Zielke HR, Zielke CL and Ozand PT (1984) Glutamine: a major energy source for cultured mammalian cells. Fed. Proc. Fed. Am. Soc. Biol. Med. 43: 121–131.
Author information
Authors and Affiliations
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 1994 Springer Science+Business Media Dordrecht
About this chapter
Cite this chapter
Butler, M., Christie, A. (1994). Adaptation of mammalian cells to non-ammoniagenic media. In: Buckland, B.C., Aunins, J.G., Bibila, T.A., Hu, WS., Robinson, D.K., Zhou, W. (eds) Cell Culture Engineering IV. Current Applications of Cell Culture Engineering, vol 1. Springer, Dordrecht. https://doi.org/10.1007/978-94-011-0257-5_10
Download citation
DOI: https://doi.org/10.1007/978-94-011-0257-5_10
Publisher Name: Springer, Dordrecht
Print ISBN: 978-94-010-4114-0
Online ISBN: 978-94-011-0257-5
eBook Packages: Springer Book Archive