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Immobilization of watermelon (Citrullus vulgaris) urease in agarose gel for urea estimation

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Abstract

Urease from dehusked seeds of watermelon was immobilized in 1.5% agarose gel with 53.9% entrapment. There was negligible leaching (<10% at 4°C) and the same gel membrane could repeatedly be used for seven days. The immobilization exhibited no apparent change in the optimum pH but there was a significant decrease in the optimum temperature (50°C as compared to 65°C for soluble urease). The immobilized urease revealed an apparentK m of 9.3±0.3 mM; 1.2 times lower than the soluble enzyme (11.4±0.2 mM). Unlike soluble enzyme which was inhibited at 200 mM urea, the immobilized urease was inhibited at 600 mM of urea and above, and about 47% activity was retained at 2 M urea. The time-dependent thermal inactivation kinetics at 48 and 52°C was found to be biphasic, in which half of the initial activity was destroyed more rapidly than the remaining half. These gel membranes were also used for estimating the urea content of the blood samples from the University hospital. The results obtained matched well with those obtained by the usual method employed in the clinical pathology laboratory. The significance of these observations is discussed.

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References

  1. George, S., M. Chellapandian, B. Sivasankar, and K. Jayaraman (1997) A new process for the treatment of fertilizer effluent using immobilized urease.Bioprocess Eng. 16: 83–85.

    Article  CAS  Google Scholar 

  2. Prakash, O., and L. S. B. Upadhyay (2005) Physicochemical characterization of watermelon urease upon immobilization in alginate beads.J. Plant Biochem. Biotechnol. 14: 209–213.

    CAS  Google Scholar 

  3. Srivastava, P. K., A. M. Kayastha, and Srinivasan (2001) Characterization of gelatin-immobilized pigconpea urease and preparation of a new urea biosensor.Biotechnol. Appl. Biochem. 34: 55–62.

    Article  CAS  Google Scholar 

  4. Dumiriu, S., M. Popa, V. Artenie, and F. Dan (1989) Bioactive polymers. 56: Urease immobilization on carboxymethylcellulose.Biotechnol. Bioeng. 34: 283–290.

    Article  Google Scholar 

  5. Reddy, K. R. C., P. K. Srivastava, P. M. Dey, and A. M. Kayastha (2004) Immobilization of pigeonpea (Cajanus cajan) urease on DEAE-cellulose paper strips for urea estimation.Biotechnol. Appl. Biochem. 39: 323–327.

    Article  Google Scholar 

  6. Onyezili, F. N. (1988) The enzyme coupling process in urease immobilization onO-alkylated nylon tubes.J. Biochem. Biophys. Methods 16: 255–262.

    Article  CAS  Google Scholar 

  7. Prakash, O., and G. Bhushan (1997) Isolation purification and partial characterization of urease from seeds of watermelon (Citrullus vulgaris).J. Plant Biochem. Biotechnol. 6: 45–47.

    CAS  Google Scholar 

  8. Prakash, O., and G. Bhushan (1998) A study of inhibition of urease from seeds of the watermelon (Citrullus vulgaris).J. Enzyme Inhib. 13: 69–77.

    Article  CAS  Google Scholar 

  9. Prakash, O., and G. Bhushan (1998) Molecular asymmetry in urease of watermelon (Citrullus vulgaris).J. Plant Biochem. Biotechnol. 7: 53–55.

    CAS  Google Scholar 

  10. Dixon, N. E., R. L. Blakeley, and B. Zerner (1980) Jack bean urease (EC 3.5.1.5) III. The involvement of active-site nickel ion in inhibition by β-mercaptoethanol, phosphoramidate, and fluoride.Can. J. Biochem. 58: 481–488.

    Article  CAS  Google Scholar 

  11. Prakash, O., and L. S. B. Upadhyay (2003) Effect of thiols of the activity of urease from dehusked seeds of water-melon (Citrullus vulgaris).Plant Sci. 164: 189–194.

    Article  CAS  Google Scholar 

  12. Lowry, O. H., N. J. Rosebrough, A. L. Farr, and R. J. Randall (1951) Protein measurement with the Folin phenol reagent.J. Biol. Chem. 193: 265–275.

    CAS  Google Scholar 

  13. Weetall, H. H., and L. S. Hersh (1969) Urease covalently coupled to porous glass.Biochim. Biophys. Acta 185: 464–465.

    CAS  Google Scholar 

  14. Iyengar, L., P. Bajpai, and A. V. Prabhakara Rao (1982) Behaviour of molecular sieve bound urease.Indian J. Biochem. Biophys. 19: 130–134.

    CAS  Google Scholar 

  15. Kayastha, A. M. and P. K. Srivastava (2001) Pigeonpea (Cajanus cajan L.) urease immobilized on glutaraldehyde-activated chitosan beads and its analytical applications.Appl. Biochem. Biotechnol. 96: 41–53.

    Article  CAS  Google Scholar 

  16. Moynihan, H. J., C. K. Lee, W. Clark and N.-H. L. Wang (1989) Urea hydrolysis by immobilized urease in a fixed-bed reactor: analysis and kinetic parameter estimation.Biotechnol. Bioeng. 34: 951–963.

    Article  CAS  Google Scholar 

  17. Guilbault, G. G., and J. Das (1970) Immobilization of cholinesterase and urease.Anal. Biochem. 33: 341–355.

    Article  CAS  Google Scholar 

  18. May, S. W., and N. N. Li (1972) The immobilization of urease using liquid-surfactant membranes.Biochem. Biophys. Res. Commun. 47: 1179–1185.

    Article  CAS  Google Scholar 

  19. Wall, M. C., and K. J. Laidler (1953) The molecular kinetics of the urea-urease system. VI. The activation of urease by amino acids.Arch. Biochem. Biophys. 43: 312–318.

    Article  CAS  Google Scholar 

  20. Malhotra, O. P. and I. Rani (1969) Purification and properties of urease ofCajanus indicus.Indian J. Biochem. 6: 15–20.

    CAS  Google Scholar 

  21. Kayastha, A. M., and N. Das (1998) Kinetics of thermal inactivation and molecular asymmetry of urease from dehusked pigeonpea (Cajanus cajan L.) seeds.J. Plant Biochem. Biotechnol. 7: 121–124.

    CAS  Google Scholar 

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Prakash, O., Puliga, S. & Bachan Upadhyay, L.S. Immobilization of watermelon (Citrullus vulgaris) urease in agarose gel for urea estimation. Biotechnol. Bioprocess Eng. 12, 131–135 (2007). https://doi.org/10.1007/BF03028638

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  • DOI: https://doi.org/10.1007/BF03028638

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