Inscrição na biblioteca: Guest
Portal Digital Begell Biblioteca digital da Begell eBooks Diários Referências e Anais Coleções de pesquisa
Catalysis in Green Chemistry and Engineering

ISSN Imprimir: 2572-9896
ISSN On-line: 2572-990X

Catalysis in Green Chemistry and Engineering

DOI: 10.1615/CatalGreenChemEng.2019029612
pages 345-355


Girish S. Nhivekar
Department of Chemical Engineering, Institute of Chemical Technology, Matunga (E), Mumbai, Maharashtra, 400019, India
Virendra K. Rathod
Department of Chemical Engineering, Institute of Chemical Technology, Matunga (E), Mumbai, 400019, India


Combi-magnetic cross-linked enzyme aggregates (CLEAs) of cellulase and lipase were successfully prepared by co-precipitation of cellulase and lipase using 80% ammonium sulphate and later cross-linking the enzyme aggregates with 100 mM glutaraldehyde cross-linker on amino-functionalized magnetic nanoparticles in 6 h. The synthesized nanoparticles and immobilized enzymes were analyzed and confirmed through X-ray diffraction and Fourier transform infrared. Various parameters affecting the immobilization such as precipitating agent, precipitant concentration, amount of cross-linker, and time of cross-linking were studied. The combi-magnetic CLEAs showed better stability at higher temperature and pH as compared to free enzymes. After immobilization, the reusability retained more than 90% for the first three cycles, which further reduced due to leaching. Vmax and Km values show that the immobilized enzyme was more active compared to the free form, while affinity of the enzyme was reduced.


  1. Aytar, B.S. and Bakir, U., Preparation of Cross-Linked Tyrosinase Aggregates, Process Biochem., vol. 43, pp. 125–131, 2008.

  2. Bajpai, P., Topical Paper, Biotechnology, Biotechnol. Prog., vol. 15, pp. 147–157, 1999.

  3. Cao, L., van Langen, L., and Sheldon, R.A., Immobilized Enzymes: Carrier-Bound or Carrier-Free? Curr. Opin. Biotechnol., vol. 14, pp. 387–394, 2003.

  4. Dalal, S., Kapoor, M., and Gupta, M.N., Preparation and Characterization of Combi-CLEAs Catalyzing Multiple Non-Cascade Reactions, J. Mol. Catal. B Enzym., vol. 44, pp. 128–132, 2007.

  5. Deshpande, A., D’souza, S.F., and Nadkarni, G.B., Coimmobilization of D-Amino Acid Oxidase and Catalase by Entrapment of Trigonopsis variabilis in Radiation Polymerised Polyacrylamide Beads, J. Biosci., vol. 11, pp. 137–144, 1987.

  6. Dyal, A., Loos, K., and Noto, M., Activity of Candida Rugosa Lipase Immobilized on &gamma;-Fe<sub>2</sub>O<sub>3</sub> Magnetic Nanoparticles, J. Am. Chem. Soc., vol. 125, pp. 1684–1685, 2003.

  7. Gu, Q.-P., You, J.-X., Yong, Q., and Yu, S.-Y., Enzymatic Deinking of ONP with Lipase/Cellulase/Xylanase, Chung-kuo Tsao Chih/China Pulp and Paper, vol. 23, pp. 7–9, 2004.

  8. Hasan, F., Shah, A.A., and Hameed, A., Industrial Applications of Microbial Lipases, Enzyme Microbe Technol., vol. 39, pp. 235– 251, 2006.

  9. Jadhav, S.B. and Singhal, R.S., Co-Conjugation vis-a-vis Individual Conjugation of &alpha;-Amylase and Glucoamylase for Hydrolysis of Starch, Carbohydr. Polym., vol. 98, pp. 1191–1197, 2013.

  10. Kar, P., Wen, H., and Li, H., Simulation of Multistep Enzyme-Catalyzed Methanol Oxidation in Biofuel Cells, J. Electrochem. Soc., vol. 158, p. B580, 2011.

  11. Karaoglu, E., Summak, M.M., and Baykal, A., Synthesis and Characterization of Catalytically Activity Fe<sub>3</sub>O<sub>4</sub>-3-Aminopropyltriethoxysilane/ Pd Nanocomposite, J. Inorg. Organomet. Polym. Mater., vol. 23, pp. 409–417, 2013.

  12. Khoshnevisan, K., Bordbar, A.K., and Zare, D., Immobilization of Cellulase Enzyme on Superparamagnetic Nanoparticles and Determination of Its Activity and Stability, Chem. Eng. J., vol. 171, pp. 669–673, 2011.

  13. Lone, M.A.,Wani, M.R., Bhat, N.A., and Sheikh, S.A., Evaluation of Cellulase Enzyme Secreted by Some Common and Stirring, Rhizoshere ungi of Juglans Regia L. by DNS Method, J. Enzyme Res., vol. 3, pp. 18–22, 2012.

  14. Lopez-Serrano, P., Cao, L., Van Rantwijk, F., and Sheldon, R.A. , Cross-Linked Enzyme Aggregates with Enhanced Activity: Application to Lipases, Biotechnol. Letters, vol. 24, no. 16, pp. 1379–1383, 2002.

  15. Minakshi Pundir, C.S., Co-Immobilization of Lipase, Glycerol Kinase, Glycerol-3-Phosphate Oxidase and Peroxidase on to Aryl Amine Glass Beads Affixed on Plastic Strip for Determination of Triglycerides in Serum, Indian J. Biochem. Biophys., vol. 45, pp. 111–115, 2008.

  16. Nadar, S.S. and Rathod, V.K., Magnetic Macromolecular Cross Linked Enzyme Aggregates (CLEAs) of Glucoamylase, Enzyme Microb. Technol., vol. 83, pp. 78–87, 2016.

  17. Pawar, S.V. and Yadav, G.D., PVA/Chitosan-Glutaraldehyde Cross-Linked Nitrile Hydratase as Reusable Biocatalyst for Conversion of Nitriles to Amides, J. Mol. Catal. B Enzyme, vol. 101, pp. 115–121, 2014.

  18. Prabhavathi Devi, B.L.A., Guo, Z., and Xu, X., Characterization of Cross-Linked Lipase Aggregates, J. Am. Oil Chem. Soc., vol. 86, pp. 637–642, 2009.

  19. Rattanachomsri, U., Tanapongpipat, S., Eurwilaichitr, L., and Champreda, V., Simultaneous Non-Thermal Saccharification of Cassava Pulp by Multi-Enzyme Activity and Ethanol Fermentation by Candida Tropicalis, J. Biosci. Bioeng., vol. 10, pp. 488– 493, 2009.

  20. Sheldon, R.A., Cross-Linked Enzyme Aggregates (CLEA®s): Stable and Recyclable Biocatalysts, Biochem. Soc. Trans., vol. 35, pp. 1583–1587, 2007.

  21. Sinirlioglu, Z.A., Sinirlioglu, D., and Akbas, F., Preparation and Characterization of Stable Cross-Linked Enzyme Aggregates of Novel Laccase Enzyme from Shewanella Putrefaciens and usingMalachite Green Decolorization, Bioresour. Technol., vol. 146, pp. 807–811, 2013.

  22. Sulaiman, N.J., Rahman, R.A., and Ngadi, N., Precipitation of Cellulase and Xylanase for Cross-Linked Enzyme Aggregates, J. Teknol. (Sciences Eng.), vol. 68, pp. 17–20, 2014.

  23. Talekar, S., Ghodake, V., and Ghotage, T., Novel Magnetic Cross-Linked Enzyme Aggregates (Magnetic CLEAs) of Alpha Amylase, Bioresour. Technol., vol. 123, pp. 542–547, 2012.

  24. Talekar, S., Joshi, A., Joshi, G., Priyanka K., Rutumbara H., and Shashikant K., Parameters in Preparation and Characterization of Cross Linked Enzyme Aggregates (CLEAs), RSC Adv., vol. 3, pp. 12485–12511, 2013.

  25. Yu, H.W., Chen, H., and Wang, X., Cross-Linked Enzyme Aggregates (CLEAs) with Controlled Particles: Application to Candida rugosa Lipase, J. Mol. Catal. B Enzym., vol. 43, pp. 124–127, 2006.

Articles with similar content:

Proceedings of the 24th National and 2nd International ISHMT-ASTFE Heat and Mass Transfer Conference (IHMTC-2017), Vol.0, 2017, issue
Khabbab Zakaria, Nurul Islam, Ranjan Ganguly, Apurba Kumar Santra, Biswaroop Roy, Pritam Kumar Das
Hepatoprotective Activity of Water Extracts from Chaga Medicinal Mushroom, Inonotus obliquus (Higher Basidiomycetes) Against Tert-Butyl Hydroperoxide−Induced Oxidative Liver Injury in Primary Cultured Rat Hepatocytes
International Journal of Medicinal Mushrooms, Vol.17, 2015, issue 11
Hyung Joo Suh, Dong Ouk Noh, Yooheon Park, Ki Bae Hong
International Journal of Energetic Materials and Chemical Propulsion, Vol.13, 2014, issue 4
Teddy Gilloux, Emilie Labarthe, Guy Jacob, Chaza Darwich, Lionel Joucla, Henri Delalu
Protective Effects of Extract from Sclerotium of the King Tuber Medicinal Mushroom, Pleurotus tuberregium (Higher Basidiomycetes) on Carbon Tetrachloride-Induced Hepatotoxicity in Wistar Albino Rats
International Journal of Medicinal Mushrooms, Vol.17, 2015, issue 12
Christopher C. Osubor, Chidube A. Alagbaoso, Omoanghe S. Isikhuemhen
Effect of Additive on the Rheological Characteristics of Slurries of Fly Ash and Bottom Ash Mixture at High Concentrations
International Journal of Fluid Mechanics Research, Vol.36, 2009, issue 6
S. N. Singh, V. Seshadri, Sunil Chandel