Journal of Thermal Analysis and Calorimetry

, Volume 93, Issue 2, pp 569–574 | Cite as

Effects of sodium phosphate buffer on horseradish peroxidase thermal stability

  • L. Haifeng
  • L. Yuwen
  • C. Xiaomin
  • W. ZhiyongEmail author
  • W. Cunxin


Thermal stability of horseradish peroxidase (HRP) was studied by differential scanning calorimetry, tryptophan fluorescence, the heme absorption and enzymatic activity analysis while the concentrations of sodium phosphate buffer ranged from 2.5 to 50 mM at pH 7.0.

The results showed that the denaturation temperature (T m) values decreased and the intrinsic tryptophan fluorescence intensity of denatured HRP increased as sodium phosphate buffer concentration increased. Furthermore, the heme absorbance at 403 nm and enzymatic activity of HRP decreased with the increasing buffer concentrations. According to data obtained in this experiment, it can be concluded that sodium phosphate accelerated the denaturation process of HRP and reduced the thermal stability of HRP.


DSC enzymatic activity fluorescence heme absorption horseradish peroxidase sodium phosphate buffer thermal stability 


Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.


  1. 1.
    H. B. Dunford, Ed., Heme Peroxidases, Wiley, New York 1999.Google Scholar
  2. 2.
    P. Tijssen and E. Kurstak, Anal. Biochem., 136 (1984) 451.CrossRefGoogle Scholar
  3. 3.
    R. Maidan and A. Heller, J. Am. Chem. Soc., 113 (1991) 9003.CrossRefGoogle Scholar
  4. 4.
    M. G. Garguilo, N. Huynh, A. Proctor and C. A. Michael, Anal. Chem., 65 (1993) 523.CrossRefGoogle Scholar
  5. 5.
    A. L. Ghindilis, P. Atanasov and E. Wilkins, Electroanalysis, 9 (1997) 661.CrossRefGoogle Scholar
  6. 6.
    M. Gajhede, D. J. Schuller, A. Henriksen, A. T. Smith and T. L. Poulos, Nat. Struct. Biol., 4 (1997) 1032.CrossRefGoogle Scholar
  7. 7.
    O. Mirza, A. Henriksen, L. Ostergaard, K. G. Welinder and M. Gajhede, Acta Crystallogr. D: Biol. Crystallogr., 56 (2000) 372.CrossRefGoogle Scholar
  8. 8.
    A. S. L. Carvalho, E. P. E. Melo, B. S. Ferreira, M. T. Neves-Petersen, S. B. Petersen and M. R. Aires-Barros, Arch. Biochem. Biophys., 415 (2003) 257.CrossRefGoogle Scholar
  9. 9.
    H. S. Pappa and A. E. G. Cass, Eur. J. Biochem., 212 (1993) 227.CrossRefGoogle Scholar
  10. 10.
    K. G. Welinder, J. M. Mauro and L. Norskov-Lauritsen, Biochem. Soc. Trans., 20 (1992) 337.Google Scholar
  11. 11.
    T. L. Poulos, Curr. Opin. Biotechnol., 4 (1993) 484.CrossRefGoogle Scholar
  12. 12.
    A. M. English and G. Tsaprailis, Adv. Inorg. Chem., 43 (1995) 79.CrossRefGoogle Scholar
  13. 13.
    A. Henriksen, O. Mirza, C. Indiani, K. Teilum, G. Smulevich, K. G. Welinder and M. Gajhede, Protein Sci., 10 (2001) 108.CrossRefGoogle Scholar
  14. 14.
    N. C. Veitch and R. J. P. Williams, Eur. J. Biochem., 189 (1990) 351.CrossRefGoogle Scholar
  15. 15.
    J. W. Tams and K. G. Welinder, Biochemistry, 35 (1996) 7573.CrossRefGoogle Scholar
  16. 16.
    G. Tsaprailis, D. W. S. Chan and A. M. English, Biochemistry, 37 (1998) 2004.CrossRefGoogle Scholar
  17. 17.
    K. Chattopadhyay and S. Mazumdar, Biochemistry, 39 (2000) 263.CrossRefGoogle Scholar
  18. 18.
    D. G. Pina, A. V. Shnyrova, F. Gavilanes, A. Rodriguez, F. Leal, M. G. Roig, I. Y. Sakharov, G. G. Zhadan, E. Villar and V. L. Shnyrov, Eur. J. Biochem., 268 (2001) 120.CrossRefGoogle Scholar
  19. 19.
    A. S. L. Carvalho, M. T. Neves-Petersen, S. B. Petersen, M. R. Aires-Barros and E. P. E. Melo, Biochim. Biophys. Acta, 1747 (2005) 99.Google Scholar
  20. 20.
    A. S. L. Carvalho, B. S. Ferreira, M. T. Neves-Petersen, S. B. Petersen, M. R. Aires-Barros and E. P. Melo, Enzyme Microb. Technol., 2006 in press.Google Scholar
  21. 21.
    G. Rialdi and E. Battistel, J. Thermal Anal., 47 (1996) 17.CrossRefGoogle Scholar
  22. 22.
    A. Ginsburg, J. Therm. Anal. Cal., 61 (2000) 425.CrossRefGoogle Scholar
  23. 23.
    G. D. Manetto, C. La Rosa, D. M. Grasso and D. Milardi, J. Therm. Anal. Cal., 80 (2005) 263.CrossRefGoogle Scholar
  24. 24.
    A. Michnik, K. Michalik and Z. Drzazga, J. Therm. Anal. Cal., 80 (2005) 399.CrossRefGoogle Scholar
  25. 25.
    B. Bugyi, G. Papp, S. Halasi and B. Visegrády, J. Therm. Anal. Cal., 82 (2005) 275.CrossRefGoogle Scholar
  26. 26.
    G. Papp, B. Bugyi, Z. Ujfalusi, S. Halasi and J. Orbán, J. Therm. Anal. Cal., 82 (2005) 281.CrossRefGoogle Scholar
  27. 27.
    A. Michnik, J. Therm. Anal. Cal., 87 (2007) 91.CrossRefGoogle Scholar
  28. 28.
    A. Stirpe, R. Guzzi, H. Wijma, M. P. Verbeet, G. W. Canters and L. Sportelli, Biochim. Biophys. Acta, 1752 (2005) 47.Google Scholar
  29. 29.
    M. Rezaei-Tavirani, S. H. Moghaddamnia, B. Ranjbar, M. Amani and S. A. Marashi, J. Biochem. Mol. Biol., 39 (2006) 530.Google Scholar
  30. 30.
    X. P. Geng, Y. N. Wu, B. H. Wang, H. F. Zhang, X. D. Geng and J. W. Xing, J. Therm. Anal. Cal., 85 (2006) 601.CrossRefGoogle Scholar
  31. 31.
    J. W. Tams and K. G. Welinder, FEBS Lett., 421 (1998) 234.CrossRefGoogle Scholar
  32. 32.
    M. A. Lemos, J. C. Oliveira and J. A. Saraiva, Lebensmittel-Wissenschaft und-Technologie, 33 (2000) 362.CrossRefGoogle Scholar
  33. 33.
    R. Lumry and E. Eyring, J. Phys. Chem., 58 (1954) 110.CrossRefGoogle Scholar
  34. 34.
    J. M. Sanchez-Ruiz, Biophys. J., 61 (1992) 921.CrossRefGoogle Scholar
  35. 35.
    I. M. Plaza del Pino, B. Ibarra-Molero and J. M. Sanchez-Ruiz, Proteins, 40 (2000) 58.CrossRefGoogle Scholar
  36. 36.
    H. F. Liu, Z.Y. Wang, Y. W. Liu, J. Xiao and C. X. Wang, Thermochim. Acta, 443 (2006) 173.CrossRefGoogle Scholar
  37. 37.
    J. E. Brunet, G. A. Gonzalez and C. P. Sotomayor, Photochem. Photobiol., 38 (1983) 253.CrossRefGoogle Scholar
  38. 38.
    T. K. Das and S. Mazumdar, Eur. J. Biochem., 227 (1995) 823.CrossRefGoogle Scholar
  39. 39.
    J. R. Lakowicz, Principles of Fluorescence Spectroscopy, Plenum Press, New York 1983.Google Scholar
  40. 40.
    A. T. Smith and N. C. Veitch, Curr. Opin. Chem. Biol., 2 (1998) 269.CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC. 2008

Authors and Affiliations

  • L. Haifeng
    • 1
  • L. Yuwen
    • 1
  • C. Xiaomin
    • 1
  • W. Zhiyong
    • 1
    Email author
  • W. Cunxin
    • 1
  1. 1.College of Chemistry and Molecular ScienceWuhan UniversityWuhan, HubeiP.R. China

Personalised recommendations