Advertisement

Bioseparation

, Volume 8, Issue 1–5, pp 247–254 | Cite as

Use of a micro-expanded bed containing immobilised lysozyme for cell disruption in flow injection analysis

  • M. P. Nandakumar
  • Anita Tocaj
  • B. Mattiasson
Article

Abstract

A method for cell disruption in Flow Injection Analysis (FIA) systems has been developed. The principle involves on-line cell disruption by means of immobilised lysozyme followed by an ultrasonic treatment. In order to avoid flow problems in the analytical system, the lysozyme was immobilised to Streamlinereg that was used in an expanded bed in the flow system. Samples of suspensions of Micrococcus lysodeikticus were treated and the success of the treatment was evaluated in terms of released protein and as a decrease in the optical density at 450 nm. The new technology offers a powerful tool in flow injection analyses for quantification of intracellular compounds. The concept of integration, i.e. combining cell disruption with handling of cell debris and assay procedure in one continuous flow process facilitates its use and increases the probability of reaching reproducible and reliable results.

cell disruption ultrasonication immobilised lysozyme flow injection analysis 

Preview

Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.

References

  1. Andrews BA and Asenjo JA (1987) Enzymatic lysis and disruption of microbial cells. Trends Biotechnol. 5: 273–277.Google Scholar
  2. Caldeira JCG and Cabral JMS (1994) Extraction of a steroid 1,2 dehydrogenase by sonication of Arthrobacter simplex cells. Bioseparation 4: 271–278.Google Scholar
  3. Chase H (1994) Purification of proteins by adsorption chromatography in expanded beds. Trends Biotechnol. 12: 296–303.Google Scholar
  4. Chen J-P and Chen Y-C (1996) Improvement of cell lysis activity of immobilised lysozyme with reversibly soluble-insoluble polymer as carrier. Biotechnol. Techniques. 10: 749–754.Google Scholar
  5. Crapisi A, Lante A, Pasini G and Spettoli P (1993) Enhanced microbial cell lysis by the use of lysozyme immobilised on different carriers. Process Biochem. 28: 17–21.Google Scholar
  6. Datta R, Armiger W and Ollis DF (1973) Lysis of Micrococcus lysodeikticus by lysozyme covalently immobilised on cellulose and polyacrylamide. Biotechnol. Bioeng. 15: 993–1006.Google Scholar
  7. Engler CD and Robinson CW (1979) New method for measuring cell-wall rupture. Biotechnol. Bioeng. 21: 1861–1869.Google Scholar
  8. Fiechter A and Sonnleitner B (1994) Non-Invasive concepts in metabolic studies. Adv. Microbial Physiol. 36: 145–180.Google Scholar
  9. Fish NM, Harbron S, Allenby DJ and Lilly MD (1983) Oxidation of n-alkenes: Isolation of alkane hydroxylase from Pseudomonas putida. Eur. J. Microbiol. Biotechnol. 17: 57–63.Google Scholar
  10. Gray PP, Dunnill P and Lilly MD (1972) Kinetics of β-galactosidase production by a constitutive mutant of Escherichia coli. J. Ferment Technol. 50: 381–387.Google Scholar
  11. Lee C-H, Tsang SK, Urakabe R and Rha CK (1979) Disintegration of dried yeast cells and its effect on protein extraction, sedimentation property, and viscosity of the cell suspension. Biotechnol. Bioeng. 21: 1–17.Google Scholar
  12. Lee C-K and Ku M-C (1994) Lysis of Micrococcus lysodeikticus cells by lysozyme covalently immobilised on the lumen of hollow fibres. Biotechnol. Techniques 8: 193–198.Google Scholar
  13. Lohmeier-Vogel E, Hahn-Hägerdal B and Vogel HJ (1986) Phosphorus-31 NMR studies on maltose and glucose metabolism in Streptococcus lactis. Appl. Microbiol. Biotechnol. 25: 43–51.Google Scholar
  14. Mattiasson B (1977) The use of coimmobilized lysozyme as a bactericide in enzyme columns: a step towards the design of self-sterilising enzyme columns. Biotechnol. Bioeng. 19: 777–780.Google Scholar
  15. Mattiasson B and Danielsson B (1996) Flow injection analysis (FIA) in combination with biosensors. In: RF Taylor and JS Schultz eds. Handbook of Chemical and Biological Sensors. IOP Publishing Ltd. Bristol and Philadelphia. pp 533–552.Google Scholar
  16. Mattiasson B and Borrebaeck C (1978) Non-equilibrium, isokinetic enzyme immunoassay of insulin using reversibly immobilised antibodies. In: B. Pal. ed. Enzyme labelled immunoassay of hormones and drugs. Walter de Gruyter & Co. New York. pp 91–105.Google Scholar
  17. Mattiasson B and Nandakumar MP (1999) Binding assays in heterogeneous media using a flow injection system with an expanded bed adsorption column. Bioseparation, this volume.Google Scholar
  18. Nakamura S, Kato A and Kobayashi K (1991) New antimicrobial characteristics of lysozyme-dextrane conjugate. J. Agric. Food Chem. 39: 647–650.Google Scholar
  19. Nandakumar MP, Lali A and Mattiasson B (1999) On-line monitoring of fermentor using expanded bed flow injection analyser. Bioseparation, this volume.Google Scholar
  20. Nilsson K and Mosbach K (1980) p-Toluenesulfonyl chloride as an activating agent of agarose for the preparation of immobilised affinity ligands and proteins. Eur. J. Biochem. 112: 397–402.Google Scholar
  21. Nilsson M, Mattiasson G and Mattiasson B (1993) Automated immunochemical binding assay (flow-ELISA) based on repeated use of an antibody column placed in a flow-injection system. J. Biotechnol. 31: 381–384.Google Scholar
  22. Ruzicka J and Hansen EH (1988) Flow injection analysis. 2nd ed. John Wiley & Sons, Inc., New York, USA.Google Scholar
  23. Sonnleitner B (1991) Dynamics of yeast metabolism and regulation. Bioproc. Eng. 6: 187–193.Google Scholar
  24. Wang DIC, Cooney CL, Demain AL, Dunhill P, Humprey AE and Lilly MD (1979) Fermentation and Enzyme Technology. John Wiley & Sons, New York. pp 238–310.Google Scholar
  25. Whitaker JR and Granum PE (1980) An absolute method for protein determination based on differences in absorbance at 235 and 280 nm. Anal. Biochem. 109: 156–159.Google Scholar

Copyright information

© Kluwer Academic Publishers 1999

Authors and Affiliations

  • M. P. Nandakumar
    • 1
  • Anita Tocaj
    • 1
  • B. Mattiasson
    • 1
  1. 1.Department of Biotechnology, Center for Chemistry and Chemical EngineeringLund UniversityLundSweden

Personalised recommendations