Skip to main content
SpringerLink
Log in

Efficient immobilization of lipase from Candida rugosa by entrapment into poly(N-isopropylacrylamide-co-itaconic acid) hydrogels under mild conditions

  • Original Paper
  • Published:
Polymer Bulletin Aims and scope Submit manuscript

Abstract

Temperature and pH-sensitive hydrogels, based on N-isopropylacrylamide and itaconic acid, with varying comonomer ratios and crosslinking agent content, were prepared by free radical crosslinking copolymerization. The immobilization of lipase from Candida rugosa was carried out by post-loading entrapment method at different temperatures until equilibrium swelling was achieved. The effects of the hydrogel composition and the immobilization temperature on the hydrogel-binding capacity, immobilized lipase specific activity, as well as the enzyme leakage were studied. It was found that the NiPAAm/IA ratio, crosslinking agent concentration and the temperature at which the entrapment was performed significantly affected the hydrogel-binding capacity. The biocatalysts obtained by entrapment into hydrogel with the highest itaconic acid content at 5 °C exhibited both highest binding capacity and the highest specific activity, but appeared to be less suitable for repeated uses than those obtained at 25 °C.

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

Access this article

Log in via an institution

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

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6
Fig. 7

Similar content being viewed by others

References

  1. Pandey A, Benjamin S, Soccol CR, Nigam P, Krieger N, Soccol VT (1999) The realm of microbial lipases in biotechnology. Biotechnol Appl Biochem 29:119–131

    CAS  Google Scholar 

  2. Paiva AL, Balcão VM, Malcata FX (2000) Kinetics and mechanisms of reactions catalyzed by immobilized lipases. Enzyme Microb Technol 27:187–204

    Article  CAS  Google Scholar 

  3. Betigeri SS, Neau SH (2002) Immobilization of lipase using hydrophilic polymers in the form of hydrogel beads. Biomaterials 23:3627–3636

    Article  CAS  Google Scholar 

  4. Ramos MC, Garcia MH, Cabral FAP, Guthrie JT (1992) Immobilization of lipase from Mucor miehei onto poly (ethylene) based graft copolymer. Biocatalysis 6:223–234

    Article  CAS  Google Scholar 

  5. Sagiroglu A, Telefoncu A (2004) Immobilization of lipases on different carriers and their use in synthesis of pentyl isovalerates. Prep Biochem Biotechnol 34:169–178

    Article  CAS  Google Scholar 

  6. Gupta S, Kumar Y, Singh K, Bhattacharya A (2010) Lipase immobilized on poly(vinyl alcohol) modified polysulfone membrane: application in hydrolytic activities for olive oil. Polym Bull 64:141–158

    Article  CAS  Google Scholar 

  7. Chang R-C, Chou S-J, Shaw J-F (2001) Synthesis of fatty acid esters by recombinant Staphylococcus epidermidis lipases in aqueous environment. J Agric Food Chem 49:2619–2622

    Article  CAS  Google Scholar 

  8. Chauhan GS, Chauhan S, Kumar Y, Thakur US, Kanwar SS, Kaushal R (2007) Designing acrylamide- and methacrylate-based novel supports for lipase immobilization. J Appl Polym Sci 105:3006–3016

    Article  CAS  Google Scholar 

  9. Tang Y, Singh J (2009) Biodegradable and biocompatible thermosensitive polymer based injectable implant for controlled release of protein. Int J Pharm 365:34–43

    Article  CAS  Google Scholar 

  10. Wang B, Zhu W, Zhang Y, Yang Z, Ding J (2006) Synthesis of a chemically-crosslinked thermo-sensitive hydrogel film and in situ encapsulation of model protein drugs. React Funct Polym 66:509–518

    Article  CAS  Google Scholar 

  11. Hoare TR, Kohane DS (2008) Hydrogels in drug delivery: progress and challenges. Polymer 49:1993–2007

    Article  CAS  Google Scholar 

  12. Milašinović N, Kalagasidis Krušić M, Knežević-Jugović Z, Filipović J (2010) Hydrogels of N-isopropylacrylamide copolymers with controlled release of a model protein. Int J Pharm 383:53–61

    Article  Google Scholar 

  13. Milašinović N, Milosavljević N, Filipović J, Knežević-Jugović Z, Kalagasidis Krušić M (2010) Synthesis, characterization and application of poly(N-isopropylacrylamide-co-itaconic acid) hydrogels as supports for lipase immobilization. React Funct Polym 70:807–814

    Article  Google Scholar 

  14. Kirimura K, Sato T, Nakanishi N, Terada M, Usami S (1997) Breeding of starch-utilizing and itaconic-acid-producing koji molds by interspecific protoplast fusion between Aspergillus terreus and Aspergillus usamii. Appl Microbiol Biotechnol 47:127–131

    Article  CAS  Google Scholar 

  15. Petruccioli M, Pulci V, Federici F (1999) Itaconic acid production by Aspergillus terreus on raw starchy materials. Lett Appl Microbiol 28:309–312

    Article  CAS  Google Scholar 

  16. Willke Th, Vorlop K-D (2001) Biotechnological production of itaconic acid. Appl Microbiol Biotechnol 56:289–295

    Article  CAS  Google Scholar 

  17. Zhang Y, Zhu W, Wang B, Ding J (2005) A novel microgel and associated post-fabrication encapsulation technique of proteins. J Control Release 105:260–268

    Article  CAS  Google Scholar 

  18. Norris RD (1975) Stabilization of aminoacetyl-tRNA synthetase by sephadex and polyacrylamide gels. Phytochemistry 14:1701–1706

    Article  CAS  Google Scholar 

  19. Cao L (2006) Enzyme entrapment. In: Cao L (ed) Carrier-bound immobilized enzymes: principles, applications and design. Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim, pp 317–395

    Chapter  Google Scholar 

  20. Weast RC (1974) Handbook of chemistry and physics 55th Edition 1974–1975. CRC Press, Cleveland OH

    Google Scholar 

  21. Hong J, Xu D, Gong P, Ma H, Dong L, Yao S (2007) Conjugation of enzyme on superparamagnetic nanogels covered with carboxyl groups. J Chromatogr B 850:499–506

    Article  CAS  Google Scholar 

  22. Peppas NA, Bures P, Leobandung W, Ichikawa H (2000) Hydrogels in pharmaceutical formulations. Eur J Pharm Biopharm 50:27–46

    Article  CAS  Google Scholar 

  23. Liu X, Fussell G, Marcolongo M, Lowman AM (2009) Characterization of associating hydrogels of poly(vinyl alcohol) and poly(vinyl pyrrolidone). J Appl Polym Sci 112:541–549

    Article  CAS  Google Scholar 

  24. Knežević Z, Milosavić N, Bezbradica D, Jakovljević Z, Prodanović R (2006) Immobilization of lipase from Candida rugosa on Eupergit® C supports by covalent attachment. Biochem Eng J 30:269–278

    Article  Google Scholar 

  25. Tietz NW, Fiereck EA (1966) A specific method for serum lipase determination. Clin Chim Acta 13:352–358

    Article  CAS  Google Scholar 

  26. Mudassir J, Ranjha NM (2008) Dynamic and equilibrium swelling studies: crosslinked pH sensitive methyl methacrylate-co-itaconic acid (MMA-co-IA) hydrogels. J Polym Res 15:195–203

    Article  CAS  Google Scholar 

  27. Aixiang Q, Mangeng LU, Qunfeng L, Ping Z (2007) Synthesis and characterization of thermo-sensitive poly (N-isopropylacrylamide) hydrogel with fast response rate. Front Chem China 2:135–139

    Article  Google Scholar 

  28. Magnin D, Dimitriu S, Magny P, Chornet E (2001) Lipase immobilization into porous chitoxan beads: activities in aqueous and organic media and lipase localization. Biotechnol Prog 17:734–741

    Article  CAS  Google Scholar 

  29. Lowman AM (2008) Smart pharmaceuticals. www.gatewaycoalition.org/files/NewEH/htmls/lowman.doc, 7 September 2008

  30. Gomes FM, Pereira EB, de Castro HF (2004) Immobilization of lipase on chitin and its use in nonconventional biocatalysis. Biomacromolecules 5:17–23

    Article  CAS  Google Scholar 

  31. Knezevic Z, Bobic S, Milutinovic A, Obradovic B, Mojovic Lj, Bugarski B (2002) Alginate-immobilized lipase by electrostatic extrusion for the purpose of palm oil hydrolysis in lecithin/isooctane system. Process Biochem 38:313–318

    Article  CAS  Google Scholar 

  32. Hung T-C, Giridhar R, Chiou S-H, Wu W-T (2003) Binary immobilization of Candida rugosa lipase on chitosan. J Mol Catal B 26:69–78

    Article  CAS  Google Scholar 

  33. Kadir S (1997) Production of biotech compounds. In: Crommelin DJA, Sindelar RD (eds) Pharmaceutical biotechnology: an introduction for pharmacists and pharmaceutical scientists. Harwood Academic Publishers, Amsterdam, pp 53–70

    Google Scholar 

  34. Korsmeyer RW, Gurny R, Doelker E, Buri P, Peppas NA (1983) Mechanisms of solute release from porous hydrophilic polymers. Int J Pharm 15:25–35

    Article  CAS  Google Scholar 

  35. Ritger PL, Peppas NA (1987) A simple equation for description of solute release. I. Fickian and non-Fickian release from non-swellable devices in form of slabs, sphere, cylinders or discs. J Control Release 5:23–36

    Article  CAS  Google Scholar 

Download references

Acknowledgments

The authors acknowledge funding from the Ministry of Education and Science of the Republic of Serbia, Project No. III 46010, “Novel encapsulation and enzyme technologies for designing of new biocatalysts and biologically active compounds targeting enhancement of food quality, safety and competitiveness” as well as Project No. 172062 “Synthesis and characterization of novel functional polymers and polymeric nanomaterials”. The authors would also like to thank Dr. Mirjana Mihajlović from the Institute for Biological Research “Siniša Stanković”, University of Belgrade, for her help in the sample lyophilization.

Author information

Authors and Affiliations

  1. Department of Organic Chemical Technology, Faculty of Technology and Metallurgy, University of Belgrade, Karnegijeva 4, 11000, Belgrade, Serbia

    Nikola Milašinović, Nedeljko Milosavljević, Jovanka Filipović & Melina Kalagasidis Krušić

  2. Department of Biochemical Engineering and Biotechnology, Faculty of Technology and Metallurgy, University of Belgrade, Karnegijeva 4, 11000, Belgrade, Serbia

    Zorica Knežević-Jugović

Authors
  1. Nikola Milašinović
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Nedeljko Milosavljević
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Jovanka Filipović
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Zorica Knežević-Jugović
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Melina Kalagasidis Krušić
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Nikola Milašinović.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Milašinović, N., Milosavljević, N., Filipović, J. et al. Efficient immobilization of lipase from Candida rugosa by entrapment into poly(N-isopropylacrylamide-co-itaconic acid) hydrogels under mild conditions. Polym. Bull. 69, 347–361 (2012). https://doi.org/10.1007/s00289-012-0737-7

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00289-012-0737-7

Keywords

Search

Navigation