Skip to main content
Log in

Lipase immobilization on magnetic microspheres via spacer arms: Effect of steric hindrance on the activity

  • Research Paper
  • Published:
Biotechnology and Bioprocess Engineering Aims and scope Submit manuscript

Abstract

Poly(styrene-acrylic acid) magnetic microspheres with an average diameter of 2 μm were successfully prepared and used as carriers to immobilize lipase. Lipase immobilized on microspheres with no spacer arm exhibited low activities, which were attributed to steric hindrance on the lipase conformation. To avoid steric effects, ethylenediamine and poly(ethylene glycol) (PEG) 400/800/4000 were utilized as spacer arms to bind the lipase to the microspheres. The immobilized lipase activities were improved using PEG 800/4000 as a spacer arm. Furthermore, the influence of enzyme loading on lipase activity was investigated, and the results indicated that enzyme overloading could exert steric effect on lipase activity. The degree of PEG modification was demonstrated to affect lipase activity because excess PEG on the surface of microspheres could interact with lipase due to its mobility, consequently reducing lipase activity.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Similar content being viewed by others

References

  1. Carrea, G. and S. Riva (2000) Properties and synthetic applications of enzymes in organic solvents. Angew. Chem. Int. Edit. 39: 2226–2254.

    Article  CAS  Google Scholar 

  2. Reetz, M. T. (2002) Lipases as practical biocatalysts. Curr. Opin. Chem. Biol. 6: 145–150.

    Article  CAS  Google Scholar 

  3. Gao, S. L., Y. J. Wang, W. W. Wang, G. S. Luo, and Y. Y. Dai (2010) Enhancing performance of lipase immobilized on methylmodified silica aerogels at the adsorption and catalysis processes: Effect of co-solvents. J. Mol. Catal. B: Enz. 62: 218–224.

    Article  CAS  Google Scholar 

  4. Chen, B., M. E. Miller, and R. A. Gross (2007) Effects of porous polystyrene resin parameters on Candida Lipase B adsorption, distribution, and polyester synthesis activity. Langmuir 23: 6467–6474.

    Article  CAS  Google Scholar 

  5. Fernandez-Lorente, G., Z. Cabrera, C. Godoy, R. Fernandez-Lafuente, J. M. Palomo, and J. M. Guisan (2008) Interfacially activated lipases against hydrophobic supports: Effect of the support nature on the biocatalytic properties. Proc. Biochem. 43: 1061–1067.

    Article  CAS  Google Scholar 

  6. Li, Y., F. Gao, W. Wei, J. B. Qu, G. H. Ma, and W. Q. Zhou (2010) Pore size of macroporous polystyrene microspheres affects lipase immobilization. J. Mol. Catal. B: Enz. 66: 182–189.

    Article  CAS  Google Scholar 

  7. Ge, J., D. N. Lu, J. Wang, and Z. Liu (2009) Lipase nanogel catalyzed transesterification in anhydrous dimethyl sulfoxide. Biomacromol. 10: 1612–1618.

    Article  CAS  Google Scholar 

  8. Wang, Y. J. and F. Caruso (2005) Mesoporous silica spheres as supports for enzyme immobilization and encapsulation. Chem. Mater. 17: 953–961.

    Article  CAS  Google Scholar 

  9. Bilkova, Z., M. Slovakova, N. Minc, C. Futterer, R. Cecal, D. Horak, M. Benes, I.I. Potier, J. Krenkova, M. Przybylski, and J. L. Viovy (2006) Functionalized magnetic micro- and nanoparticles: Optimization and application to μ-chip tryptic digestion. Electrophor. 27: 1811–1824.

    Article  CAS  Google Scholar 

  10. Bruice, T. C. and P. Y. Bruice (2005) Covalent intermediates and enzyme proficiency. J. Am. Chem. Soc. 127: 12478–12479.

    Article  CAS  Google Scholar 

  11. Carolan, N., R. J. Forster, and C. O. Fagain (2007) Covalent attachment of ferrocene to soybean peroxidase glycans: Electron transfer mediation to redox enzymes. Bioconjugate Chem. 18: 524–529.

    Article  CAS  Google Scholar 

  12. Chiou, S. H. and W. T. Wu (2004) Immobilization of Candida rugosa lipase on chitosan with activation of the hydroxyl groups. Biomaterials 25: 197–204.

    Article  CAS  Google Scholar 

  13. Knezevic, Z., N. Milosavic, D. Bezbradica, Z. Jakovljevic, and R. Prodanovic (2006) Immobilization of lipase from Candida rugosa on Eupergit® supports by covalent attachment. Biochem. Eng. J. 30: 269–278.

    Article  CAS  Google Scholar 

  14. Das, R. D., S. Maji, S. Das, and C. R. Chaudhuri (2010) Optimization of covalent antibody immobilization on macroporous silicon solid supports. Appl. Surf. Sci. 256: 5867–5875.

    Article  CAS  Google Scholar 

  15. Tsai, H. C. and R. A. Doong (2007) Preparation and characterization of urease-encapsulated biosensors in poly (vinyl alcohol)-modified silica sol-gel materials. Biosens. Bioelectron. 23: 66–73.

    Article  CAS  Google Scholar 

  16. Tischer, W. and V. Kasche (1999) Immobilized enzymes: Crystals or carriers? Trends Biotechnol. 17: 326–335.

    Article  CAS  Google Scholar 

  17. Zhang, D.-H., L.-X. Yuwen, C. Li, and Y.-Q. Li (2012) Effect of poly(vinyl acetate-acrylamide) microspheres properties and steric hindrance on the immobilization of Candida rugosa lipase. Bioresour. Technol. 124: 233–236.

    Article  CAS  Google Scholar 

  18. Wang, F., Z Gu, Z. Cui, and L. Liu (2011) Comparison of covalent immobilization of amylase on polystyrene pellets with pentaethylenehexamine and pentaethylene glycol spacers. Bioresour. Technol. 102: 9374–9379.

    Article  CAS  Google Scholar 

  19. Hinterwirth, H., W. Lindner, and M. Lammerhofer (2012) Bioconjugation of trypsin onto gold nanoparticles: Effect of surface chemistry on bioactivity. Anal. Chim. Acta 733: 90–97.

    Article  CAS  Google Scholar 

  20. Manta, C., N. Ferraz, L. Betancor, G. Antunes, F. Batista-Viera, J. Carlsson, and K. Caldwell (2003) Polyethylene glycol as a spacer for solid-phase enzyme immobilization. Enz. Microb. Technol. 33: 890–898.

    Article  CAS  Google Scholar 

  21. Zhang, D.-H., L.-X. Yuwen, Y.-L. Xie, W. Li, and X.-B. Li (2012) Improving immobilization of lipase onto magnetic microspheres with moderate hydrophobicity/hydrophilicity. Colloid. Surface. B. 89: 73–78.

    Article  CAS  Google Scholar 

  22. Bradford, M. M. (1976) A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal. Biochem. 72: 248–254.

    Article  CAS  Google Scholar 

  23. Yong, Y., Y. X. Bai, Y. F. Li, L. Lin, Y. J. Cui, and C. G. Xia (2008) Characterization of Candida rugosa lipase immobilized onto magnetic microspheres with hydrophilicity. Proc. Biochem. 43: 1179–1185.

    Article  CAS  Google Scholar 

  24. El-Sherif, H., P. L. Martelli, R. Casadio, M. Portaccio, U. Bencivenga, and D. G. Mita (2001) Urease immobilization on chemically grafted nylon membranes Part 1: Isothermal characterization. J. Mol. Catal. B: Enz. 14: 15–29.

    Article  CAS  Google Scholar 

  25. Bhardwaj, A., J. Lee, K. Glauner, S. Ganapathi, D. Bhattacharyya, and D. A. Butterfield (1996) Biofunctional membranes: An EPR study of active site structure and stability of papain noncovalently immobilized on the surface of modified poly(ether) sulfone membranes through the avidin-biotin linkage. J. Membrane Sci. 119: 241–252.

    Article  CAS  Google Scholar 

Download references

Author information

Authors and Affiliations

Authors

Corresponding author

Correspondence to Dong-Hao Zhang.

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Zhang, DH., Li, YQ., Peng, LJ. et al. Lipase immobilization on magnetic microspheres via spacer arms: Effect of steric hindrance on the activity. Biotechnol Bioproc E 19, 838–843 (2014). https://doi.org/10.1007/s12257-013-0495-x

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s12257-013-0495-x

Keywords

Navigation