3 Biotech

, 8:288 | Cite as

Extractive purification of recombinant thermostable lipase from fermentation broth of Escherichia coli using an aqueous polyethylene glycol impregnated resin system

  • Joo Shun TanEmail author
  • Sahar Abbasiliasi
  • Arbakariya B. Ariff
  • Hui Suan Ng
  • Mohamad Hafizi Abu Bakar
  • Yin Hui Chow
Original Article


This study aimed at recovery of thermostable lipase from Escherichia coli BL21 using porous glass beads grafted with polyethylene glycol (PEG) in aqueous impregnated resins system (AIRS). The influencing parameters such as concentration and pH of extraction solution, concentration of NaCl, size of the beads, and pH of the desorption solution on the partition behaviour of lipase were evaluated. Smaller adsorbent (4 mm) had a 65.5% of recovery yield with approximately two-fold higher purification factor compared to that obtained with the larger adsorbent. Recombinant lipase was purified successfully using AIRS with a purification factor of 7.6 and yield of 78.4% under optimum conditions of 18% (w/w) PEG 4000, 10% (w/w) of potassium citrate at pH 9 with 3% (w/w) of NaCl. Optimum desorption was obtained with 4.0 mm of porous glass beads at pH 9.


Aqueous impregnated resins system Aqueous phase Purification Recovery Lipase 



This work was funded by Universiti Sains Malaysia under USM Short Term Grant (PO4865, 304/PTENKIND/6315138).

Compliance with ethical standards

Conflict of interest

The authors declare that they have no competing interests.


  1. Anvari M (2015) Extraction of lipase from Rhizopus microsporus fermentation culture by aqueous two-phase partitioning. Biotechnol Biotechnol Equip 29(4):723–731CrossRefGoogle Scholar
  2. Aziz NFHA, Abbasiliasi S, Ng HS, Phapugrangkul P, Bakar MHA, Tam YJ, Tan JS (2017) Purification of β-mannanase derived from Bacillus subtilis ATCC 11774 using ionic liquid as adjuvant in aqueous two-phase system. J Chromatogr B 1055:104–112CrossRefGoogle Scholar
  3. Babic K (2008) Extractant impregnated resins for the recovery of aldehydes and chiral resolution. University of Twente, EnschedeCrossRefGoogle Scholar
  4. Bradford MM (1976) A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem 72(1–2):248–254CrossRefGoogle Scholar
  5. Chen J, Sun Y (2003) Modeling of the salt effects on hydrophobic adsorption equilibrium of protein. J Chromatogr A 992(1):29–40CrossRefPubMedGoogle Scholar
  6. Gedikli M, Ceylan Ş, Erzengin M, Odabaşı M (2014) A novel matrix for hydrophobic interaction chromatography and its application in lysozyme adsorption. Acta Biochim Pol 61(4):731–737PubMedGoogle Scholar
  7. González-Valdez J, Cueto LF, Benavides J, Rito-Palomares M (2011) Potential application of aqueous two-phase systems for the fractionation of RNase A and α-lactalbumin from their PEGylated conjugates. J Chem Technol Biotechnol 86(1):26–33CrossRefGoogle Scholar
  8. Gupta N, Rathi P, Singh R, Goswami VK, Gupta R (2005) Single-step purification of lipase from Burkholderia multivorans using polypropylene matrix. Appl Microbiol Biotechnol 67(5):648–653CrossRefPubMedGoogle Scholar
  9. Hadzir MH, Abbasiliasi S, Ariff AB, Yusoff SB, Ng HS, Tan JS (2016) Partitioning behavior of recombinant lipase in Escherichia coli by ionic liquid-based aqueous two-phase systems. RSC Adv 6(86):82571–82580CrossRefGoogle Scholar
  10. Iqbal M, Tao Y, Xie S, Zhu Y, Chen D, Wang X, Huang L, Peng D, Sattar A, Shabbir MAB (2016) Aqueous two-phase system (ATPS): an overview and advances in its applications. Biol Proced Online 18(1):18CrossRefPubMedPubMedCentralGoogle Scholar
  11. Ketnawa S, Rungraeng N, Rawdkuen S (2017) Phase partitioning for enzyme separation: an overview and recent applications. Int Food Res J 24(1):1–24Google Scholar
  12. Kumar A, Dhar K, Kanwar SS, Arora PK (2016) Lipase catalysis in organic solvents: advantages and applications. Biol Proced Online 18(1):2CrossRefPubMedPubMedCentralGoogle Scholar
  13. Nandini K, Rastogi NK (2010) Separation and purification of lipase using reverse micellar extraction: optimization of conditions by response surface methodology. Biotechnol Bioprocess Eng 15(2):349–358CrossRefGoogle Scholar
  14. Ooi CW, Tey BT, Hii SL, Ariff A, Wu HS, Lan JCW, Juang RS, Kamal SMM, Ling TC (2009) Direct purification of Burkholderia pseudomallei lipase from fermentation broth using aqueous two-phase systems. Biotechnol Bioprocess Eng 14(6):811–818CrossRefGoogle Scholar
  15. Oscarsson S, Kårsnäs P (1998) Salt-promoted adsorption of proteins onto amphiphilic agarose-based adsorbents: II. Effects of salt and salt concentration 1. The raw data table can be obtained from the authors upon request. J Chromatogr A 803(1):83–93CrossRefPubMedGoogle Scholar
  16. Queiroz J, Tomaz C, Cabral J (2001) Hydrophobic interaction chromatography of proteins. J Biotechnol 87(2):143–159CrossRefPubMedGoogle Scholar
  17. Ramyadevi D, Subathira A, Saravanan S (2012) Aqueous two-phase poly (ethylene glycol)–Maltodextrin system for protein partitioning from shrimp waste: influence of molecular weight and pH. Int J Environ Sci 2(4):2462–2469Google Scholar
  18. Tan JS, Abbasiliasi S, Lin YK, Mohamed MS, Kapri MR, Kadkhodaei S, Tam YJ, Rahman RNZRA, Ariff AB (2014) Primary recovery of thermostable lipase 42 derived from recombinant Escherichia coli BL21 in aqueous two-phase flotation. Sep Purif Technol 133:328–334CrossRefGoogle Scholar
  19. Tan CH, Show PL, Ooi CW, Ng EP, Lan JCW, Ling TC (2015) Novel lipase purification methods—a review of the latest developments. Biotechnol J 10(1):31–44CrossRefPubMedGoogle Scholar
  20. van Winssen F, Merz J, Czerwonka L-M, Schembecker G, Dortmund T (2014a) Application of the tunable aqueous polymer-phase impregnated resins-technology for protein purification. Sep Purif Technol 136:123–129CrossRefGoogle Scholar
  21. van Winssen F, Merz J, Schembecker G (2014b) Tunable aqueous polymer-phase impregnated resins-technology—a novel approach to aqueous two-phase extraction. J Chromatogr A 1329:38–44CrossRefPubMedGoogle Scholar
  22. Shabram P, Vellekamp G, Ruan Q, Scandella C (2002) Purification of adenovirus. In: Curiel DT (ed) Adenoviral vectors for gene therapy, 2nd edn. Academic Press, San Diego, pp 197–230Google Scholar
  23. Wu J, Zhao C, Lin W, Hu R, Wang Q, Chen H, Li L, Chen S, Zheng J (2014) Binding characteristics between polyethylene glycol (PEG) and proteins in aqueous solution. J Mater Chem B 2(20):2983–2992CrossRefGoogle Scholar
  24. Zafarani-Moattar MT, Hamzehzadeh S (2011) Effect of pH on the phase separation in the ternary aqueous system containing the hydrophilic ionic liquid 1-butyl-3-methylimidazolium bromide and the kosmotropic salt potassium citrate at T = 298.15 K. Fluid Phase Equilib 304(1):110–120CrossRefGoogle Scholar

Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2018

Authors and Affiliations

  • Joo Shun Tan
    • 1
    Email author
  • Sahar Abbasiliasi
    • 2
    • 3
  • Arbakariya B. Ariff
    • 4
  • Hui Suan Ng
    • 5
  • Mohamad Hafizi Abu Bakar
    • 1
  • Yin Hui Chow
    • 6
  1. 1.Bioprocess Technology, School of Industrial TechnologyUniversiti Sains MalaysiaGelugorMalaysia
  2. 2.Department of Microbiology, Faculty of Biotechnology and Biomolecular SciencesUniversiti Putra MalaysiaSerdangMalaysia
  3. 3.Halal Products Research InstituteUniversiti Putra MalaysiaSerdangMalaysia
  4. 4.Department of Bioprocess Technology, Faculty of Biotechnology and Biomolecular SciencesUniversiti Putra MalaysiaSerdangMalaysia
  5. 5.Department of Food Science and Nutrition, Faculty of Applied SciencesUCSI UniversityKuala LumpurMalaysia
  6. 6.School of EngineeringTaylor’s UniversitySubang JayaMalaysia

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