Acta Alimentaria
Volume/Issue:
Volume 39: Issue 3
Immobilization of polygalacturonase from Aspergilus niger onto glutaraldehyde activated Nylon-6 and its application in apple juice clarification
Authors: S. Shukla 1 , S. Saxena 1 , J. Thakur 1 , and R. Gupta 1
View More View Less
  • 1 Himachal Pradesh University Department of Biotechnology Summer Hill Shimla 171 005 India
DOI:
https://doi.org/10.1556/aalim.39.2010.3.4
Page Count:
277–292
Publication Date:
01 Sep 2010
Online Publication Date:
15 Sep 2010
Article Category:
Research Article
Restricted access

Purchase article

USD  $25.00

1 year subscription (Individual Only)

USD  $878.00
Purchase Online Subscription

  • Purchase article

    USD  $25.00

  • 1 year subscription (Individual Only)

    USD  $878.00

The application of immobilized enzyme for catalyzing various biotransformation processes is a widely used approach at present. This work mainly focused on the immobilization of polygalacturonase from Aspergillus niger Van Tieghem (MTCC 3323) on Nylon-6 by covalent binding, keeping in view its applicability in apple juice clarification. The immobilized enzyme was characterized in terms of kinetic parameters, thermal stability and reusability. The enzyme was immobilized onto glutaraldehyde-activated Nylon-6 by covalent binding and the efficiency of immobilization was found to be 40%. The immobilization yielded a protein loading of 70 μg g −1 of Nylon-6. Immobilized enzyme showed maximum activity at a temperature of 50 °C and pH 5.0. The enzyme was stable between pH 4.0–5.5. The immobilized enzyme could be reused through 4 cycles with almost 50% retention of its original activity. It had increased thermostability over its soluble form at 25 °C and 45 °C. Kinetic parameters Km and Vmax were found to be 7.6 mg ml −1 and 41.66 μmol of galacturonic acid/ml/min, respectively. The immobilized enzyme when used for apple juice clarification showed about 50% increase in transmittance of apple juice at 650 nm. This increase was observed at enzyme concentration of 20 U ml −1 apple juice, temperature 50 °C and incubation time of one hour. The optimization of these factors, which affect the stability and productivity of the immobilized system, resulted in an increase in enzyme stability and the possibility of economic application of immobilized enzyme at large scale apple juice clarification.

  • Alkorta, I., Garbisu, C., Llama, M.J. & Serra, J.L. (1996): Immobilization of pectin lyase from Penicillium italicum by covalent bonding to nylon. Enzyme Microbial Technol. , 18 , 141–146.

    Serra J.L. , 'Immobilization of pectin lyase from Penicillium italicum by covalent bonding to nylon ' (1996 ) 18 Enzyme Microbial Technol. : 141 -146.

    • Search Google Scholar

  • Findlay, C.J., Parkin, K.L. & Yada, R.Y. (1986): Bone as a solid support for the immobilization of enzymes. Biotechnol. Lett. , 8 , 649–652.

    Yada R.Y. , 'Bone as a solid support for the immobilization of enzymes ' (1986 ) 8 Biotechnol. Lett. : 649 -652.

    • Search Google Scholar

  • Fogarty, W.M. & Kelly, C.T. (1983): Pectic enzymes. -in: Fogarty, W.M. (Ed.) Microbial enzymes and biotechnology . Applied Science Publishers, London, pp. 131–182.

    Kelly C.T. , '', in Microbial enzymes and biotechnology , (1983 ) -.

  • Gadre, R.V., Driessche, G.V., Beeumen, J.V. & Bhat, M.K. (2003): Purification, characterization and mode of action of an endo-polygalacturonase from the psychrophilic fungus Mucor flavus . Enzyme Microbial Technol. , 32 , 321–330.

    Bhat M.K. , 'Purification, characterization and mode of action of an endo-polygalacturonase from the psychrophilic fungus Mucor flavus ' (2003 ) 32 Enzyme Microbial Technol. : 321 -330.

    • Search Google Scholar

  • Gupta, R., Kumar, A., Kumar, N. & Bhalla, T.C. (2003): Polygalacturonase from Aspergillus niger : Optimization of production conditions and its application. -in: Marwaha, S.S. & Arora, J. K. (Eds) Biotechnological strategies in agroprocessing . Asiatech Publishers Inc, New Delhi, India, pp. 395–400.

    Bhalla T.C. , '', in Biotechnological strategies in agroprocessing , (2003 ) -.

  • Jayani, R.S., Saxena, S. & Gupta, R. (2005): Microbial pectinolytic enzymes: A review. Process Biochem. , 40 , 2931–2944.

    Gupta R. , 'Microbial pectinolytic enzymes: A review ' (2005 ) 40 Process Biochem. : 2931 -2944.

    • Search Google Scholar

  • Lee, W.C., Yusof, S., Hamid, S.A. & Baharin, B.S. (2006): Optimizing conditions for enzymatic clarification of banana juice using response surface methodology (RSM). J. Fd Engng , 73 , 55–63.

    Baharin B.S. , 'Optimizing conditions for enzymatic clarification of banana juice using response surface methodology (RSM) ' (2006 ) 73 J. Fd Engng : 55 -63.

    • Search Google Scholar

  • Lowry, O.H., Rosebrough, N.S., Farr, A.L. & Randall, R.J. (1951): Protein measurement with the Folin phenol reagent. J. biol. Chem. , 193 , 265–275.

    Randall R.J. , 'Protein measurement with the Folin phenol reagent ' (1951 ) 193 J. biol. Chem. : 265 -275.

    • Search Google Scholar

  • Lozano, P., Manjon A., Romojaro, F. & Iborra, J.L. (1987): Activity of pectolytic enzymes immobilized to nylon for viscous juice clarification. Dependence on covalent attachment method. -in: Neijssel, O.M., Van Der Meer, R.R. & Luyben, K.Ch.A.M. (Eds) Proceedings of the Fourth European Congress of Biotechnology . Vol 2. Elsevier Science, Amsterdam, pp. 52–55.

    Iborra J.L. , '', in Proceedings of the Fourth European Congress of Biotechnology. Vol 2 , (1987 ) -.

  • Lozano, P., Manjon, A., Iborra, J.L. & Galvez, D.M. (1990): Characteristics of the immobilized pectin lyase activity from a commercial pectolytic enzyme preparation. Acta biotechnol. , 10 , 531–539.

    Galvez D.M. , 'Characteristics of the immobilized pectin lyase activity from a commercial pectolytic enzyme preparation ' (1990 ) 10 Acta biotechnol. : 531 -539.

    • Search Google Scholar

  • Nelson, N. (1944): A photometric adaptation of the Somogyi method for the determination of glucose. J. biol. Chem. , 153 , 375–380.

    Nelson N. , 'A photometric adaptation of the Somogyi method for the determination of glucose ' (1944 ) 153 J. biol. Chem. : 375 -380.

    • Search Google Scholar

  • Omelkova, J., Rexova-Benkova, L., Kubanek, V. & Veruovic, B. (1985): Endopolygalacturonase immobilized on a porous poly (6-caprolactam). Biotechnol. Lett. , 7 , 99–104.

    Veruovic B. , 'Endopolygalacturonase immobilized on a porous poly (6-caprolactam) ' (1985 ) 7 Biotechnol. Lett. : 99 -104.

    • Search Google Scholar

  • Rao, M.N., Kembhavi, A.A. & Pant, A. (2000): Immobilization of endo-polygalacturonase from Aspergillus ustus on silica gel. Biotechnol. Lett. , 22 , 1557–1559.

    Pant A. , 'Immobilization of endo-polygalacturonase from Aspergillus ustus on silica gel ' (2000 ) 22 Biotechnol. Lett. : 1557 -1559.

    • Search Google Scholar

  • Rexova-Benkova, L. & Mrackova-Dobrotova, M. (1981): Effect of immobilization of Aspergillus niger extracellular endo-D-galacturonase on kinetics and action pattern. Carbohydr. Res. , 98 , 115–122.

    Mrackova-Dobrotova M. , 'Effect of immobilization of Aspergillus niger extracellular endo-D-galacturonase on kinetics and action pattern ' (1981 ) 98 Carbohydr. Res. : 115 -122.

    • Search Google Scholar

  • Rexova-Benkova, L., Omelkova, J. & Kubanek, V. (1982): Endo-D-galacturonase immobilized by adsorption on porous polyethylene terephthalate. Coll. Czech Chem. Commun. , 47 , 2716–2723.

    Kubanek V. , 'Endo-D-galacturonase immobilized by adsorption on porous polyethylene terephthalate ' (1982 ) 47 Coll. Czech Chem. Commun. : 2716 -2723.

    • Search Google Scholar

  • Rexova-Benkova, L., Omelkova, J., Veruovic, B. & Kubanek, V. (1983): Endopolygalacturonase immobilized on a porous poly (2,6-dimethyl- p -phenylene oxide). Biotechnol. Lett. , 5 , 737–742.

    Kubanek V. , 'Endopolygalacturonase immobilized on a porous poly (2,6-dimethyl-p-phenylene oxide) ' (1983 ) 5 Biotechnol. Lett. : 737 -742.

    • Search Google Scholar

  • Rexova-Benkova, L., Omelkova, J., Veruovic, B. & Kubanek, V. (1989): Mode of action endopolygalacturonase immobilized by adsorption and by covalent binding via amino groups. Biotechnol. Bioengng. , 34 , 79–85.

    Kubanek V. , 'Mode of action endopolygalacturonase immobilized by adsorption and by covalent binding via amino groups ' (1989 ) 34 Biotechnol. Bioengng. : 79 -85.

    • Search Google Scholar

  • Romero, C., Manjon, A. & Iborra, J.L. (1987): Analysis of pectinmethylesterase activity when co-immobilized with polygalacturonase on a porous glass support. -in: Neijsel, O.M., Van Der Meer, R.R. & Luyben, K.Ch.A.M. (Eds) Proceedings of the Fourth European Congress of Biotechnology . Vol. 2. Elsevier Science, Amsterdam, pp. 48–51.

    Iborra J.L. , '', in Proceedings of the Fourth European Congress of Biotechnology. Vol. 2 , (1987 ) -.

  • Sardar, M., Ro, I. & Gupta, N. (2003): A smart bioconjugate of alginate and pectinase with unusual activity toward chitosan. Biotechnol. Progress , 19 , 1654–1658.

    Gupta N. , 'A smart bioconjugate of alginate and pectinase with unusual activity toward chitosan ' (2003 ) 19 Biotechnol. Progress : 1654 -1658.

    • Search Google Scholar

  • Saxena, S., Shukla, S. & Gupta, R. (2008): Immobilization of polygalacturonase from Aspergillus niger on to activated polyethylene and its application in apple juice clarification. Acta microbiol. immunol. Hung. , 55 , 33–51.

    Gupta R. , 'Immobilization of polygalacturonase from Aspergillus niger on to activated polyethylene and its application in apple juice clarification ' (2008 ) 55 Acta microbiol. immunol. Hung. : 33 -51.

    • Search Google Scholar

  • Sin, H.N., Yusof, S., Hamid, S.A. & Rahman, R.A. (2006): Optimization of enzymatic clarification of sapodilla juice using response surface methodology. J Fd Engng , 73 , 313–319.

    Rahman R.A. , 'Optimization of enzymatic clarification of sapodilla juice using response surface methodology ' (2006 ) 73 J Fd Engng : 313 -319.

    • Search Google Scholar

  • Singh, S. & Gupta, R. (2004): Apple juice clarification using fungal pectinolytic enzyme and gelatine. Indian J. Biotechnol. , 3 , 573–576.

    Gupta R. , 'Apple juice clarification using fungal pectinolytic enzyme and gelatine ' (2004 ) 3 Indian J. Biotechnol. : 573 -576.

    • Search Google Scholar

  • Singh, S.A. & Rao, A.G.A. (2002): A simple fractionation protocol for, and a comprehensive study of the molecular properties of two major endo-polygalacturonases from Aspergillus niger . Biotechnol. appl. Biochem. , 35 , 115–123.

    Rao A.G.A. , 'A simple fractionation protocol for, and a comprehensive study of the molecular properties of two major endo-polygalacturonases from Aspergillus niger ' (2002 ) 35 Biotechnol. appl. Biochem. : 115 -123.

    • Search Google Scholar

  • Somogyi, M. (1952): Notes on sugar determination. J. biol. Chem. , 195 , 19–25.

    Somogyi M. , 'Notes on sugar determination ' (1952 ) 195 J. biol. Chem. : 19 -25.

  • Souza, J.V.B., Silva, T.M., Maia, M.L.S. & Teixeira, M.F.S. (2003): Screening of fungal strains for pectinolytic activity: endopolygalacturonase production by Peacilomyces clavisporus 2A.UMIDA.1. Process Biochem. , 4 , 455–458.

    Teixeira M.F.S. , 'Screening of fungal strains for pectinolytic activity: endopolygalacturonase production by Peacilomyces clavisporus 2A.UMIDA.1 ' (2003 ) 4 Process Biochem. : 455 -458.

    • Search Google Scholar

  • Sundaram, P.V. & Hornby, W.E. (1970): Preparation and properties of urease chemically attached to Nylon tube. FEBS Letters , 10 , 325–332.

    Hornby W.E. , 'Preparation and properties of urease chemically attached to Nylon tube ' (1970 ) 10 FEBS Letters : 325 -332.

    • Search Google Scholar

  • Whitaker, J.R. (1990): New and future uses of enzymes in food processing. Fd Biotechnol. , 4 , 669–697.

    Whitaker J.R. , 'New and future uses of enzymes in food processing ' (1990 ) 4 Fd Biotechnol. : 669 -697.

    • Search Google Scholar

  • Zaidi, A., Grainer, J.L. & Carta, G. (1995): Fatty acid esterification using nylon-immobilized lipase. Biotechnol. Bioengng , 48 , 601–605.

    Carta G. , 'Fatty acid esterification using nylon-immobilized lipase ' (1995 ) 48 Biotechnol. Bioengng : 601 -605.

    • Search Google Scholar
Cover Acta Alimentaria
Acta Alimentaria
Print ISSN:
0139-3006
Online ISSN:
1588-2535
Keywords:
Aspergillus niger ; polygalacturonase; immobilization; Nylon-6

Monthly Content Usage

Abstract Views Full Text Views PDF Downloads
Aug 2020 1 0 0
Sep 2020 4 0 0
Oct 2020 0 0 0
Nov 2020 3 1 0
Dec 2020 12 0 0
Jan 2021 0 0 0
Feb 2021 0 0 0