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İmmobilize ve Serbest Lakkaz Enziminin Optimum Koşullarının Araştırılması ve Renk Giderme İşleminde Kullanımı

Year 2019, Volume: 11 Issue: 1, 153 - 162, 31.01.2019
https://doi.org/10.29137/umagd.508095

Abstract

Bu çalışmada Tramates versicolor dan elde edilen Laccase
enzimi (L), poli(N-izopropilakrilamit)-kalsiyum aljinat (P(NIPA)-CaAlj) taneciklerine
hapsetme yöntemi ile immobilize edildi. İmmobilize enzimin özellikleri ve enzimatik
renk giderme işlemlerinde kullanımı araştırıldı ve serbest enzim ile
karşılaştırıldı.  Michaelis-Menten sabiti
(Km) ve maksimum reaksiyon hızı (Vmax) değerleri sırasıyla serbest enzim için
1,70x10-2 mM ve 2,08x10-3 mM.dak-1 olarak
bulundu. İmmobilize enzim için Km ve Vmax değerleri de sırasıyla 4,80x10-2
mM ve 8,70 x 10-3 mM.dak-1 olarak bulundu. Optimum
pH değerleri serbest enzim için 5,0 ve immobilize enzim için 6,0 olarak
belirlendi. Optimum sıcaklık sırasıyla serbest lakkaz ve immobilize lakkaz için
40oC ve 45oC olarak belirlendi. 4 oC tutulan
serbest lakkazın 30 günlük depolama sonrasında orijinal aktivitesinin % 60'ı
koruduğu bulunurken aynı koşullarda tutulan immobilize enzimin ise orijinal
aktivitesinin% 83’ ünü koruduğu bulundu. İmmobilize enzimin 10 kez tekrar
kullanım sonrasında orijinal aktivitesinin % 77'sini koruduğu bulunmuştur. Metil
oranjın renginin giderilmesinde, serbest lakkaz ve immobilize lakkaz için renk
giderme yüzdeleri sırasıyla %73 ve %70 olarak bulundu

References

  • Al-Adhami, A.J.H., Bryjak, J., Markiewicz, B.G., Chozch, W.P., (2002). Immobilization of wood-rotting fungi laccases on modified cellulose and acrylic carriers. Process Biochem, 37: 1387-1394.
  • Bamforth, S.M., Singleton, I., (2005). Bioremediation of polycyclic aromatic hydrocarbons: current knowledge and future directions. J Chem Technol Biotechnol 80: 723–736.
  • Curulli A., Cusma A., Kaciulis S., Padeletti G., Pandolfi L., Valentini F., Vitocelli M., (2006). Immobilization of GOD and HRP enzyme on nanostructured substrates. Surf. Interface Anal., 38 478–481.
  • D’Annibale, A., Stazi, A., Vinciguerra, S.R. and Giovannozzi, V.G., (2000). Oxirane-immobilized Lentinula edodes laccase: stability and phenolics removal efficiency in olive mill wastewater. Biotechnol, 77: 265-273.
  • Davis, S., and Burns, R. G. (1990). Decolorization of phenolic effluents by soluble and immobilized phenol oxidases. Appl. Microbiol. Biotechnol. 32, 721–726.
  • Davis, S., and Burns, R. G. (1992). Covalent immobilization of laccase on activated carbon for phenolic effluent treatment. Appl. Microbiol. Biotechnol. 37, 474–479.
  • Fang, H., Huang, J., Ding, L., Li, M., Chen, Z., (2009). Preparation of magnetic Chitosan nanoparticles and immobilization of Laccase. Journal of Wuhan University of Technology-Mater. Sci. Ed.Feb. 24 (1).
  • Freire, R.S., Durán, N., Kubota, L.T., (2001). Effects of fungal laccase immobilization procedures for the development of a biosensor for phenol. Talanta, 54: 681-686.
  • Gianfreda, L., Xu, F., and Bollag, J. M. (1999). Laccases: A Useful Group of Oxidoreductive Enzymes. Bioremediat. J. 3, 1–26.
  • Gokgoz M. and Altinok H., (2012). Immobilization of laccase on polyacrylamide and polyacrylamide - κ - carragennan-based semi-interpenetrating polymer networks. Artificial Cells, Blood Substitutes, and Biotechnology, 40 326–330.
  • Gomez, J., Pazos, M., Couto,R., Sanroman, M., (2005). Chestnut shell and barley bran as potential substrates for lacasse production by Corilopsis rigida under solid-state conditions. Journal of Food Engineering, 68: 315-319.
  • Hu, X., Zhau, X. and Hwang, H., (2007). Comparative study of immobilized Trametes versicolor laccase on nonoparticles and kaolinite. Chemosphere, 66: 1618-1626.
  • Hublik, G. and Schinner, F., (2000). Characterization and immobilization of the Laccase from Pleuratus ostreatus and its use for the continuous elimination of fenolic pollutants. Enzyme Microb Tech, 27: 330-336.,
  • Leonowicz, A., Grzywnowicz, K., (1981). Quantitative estimation of laccase forms in some white-rot fungi using syringaldazine as a substrate. Enzyme Microb Tech, 3: 55-58.
  • Luterek, J., Gianfreda, L., Wojtas-Wasilewska, M., Cho, N., Rogalski, J.,Jaszek, M.,Malarczyk, E.,Staszczak, M.,Fink Boots, M., Leonowicz, A., (1998). Activity of free and immobilized extracellular Cerrena unicolor Laccase in water miscible organic solvents. Cat. Inist, 52: 589-595.
  • Makas Y.G. , Kalkan N.A., Aksoy S., Altinok H., Hasirci N., (2010). Immobilization of laccase in -carrageenan based semi-interpenetrating polymer Networks. Journal of Biotechnology, 148 216–220.
  • Milstein, O., Haars, A., Majerczyck, A., Trojanowski, J., Tautz, D., Zanker, H., Hu¨ ttermann, A. (1988). Removal of Chlorophenols and Chlorolignins from Bleaching Effluent by Combined Chemical and Biological Treatment. Water Sci. Technol. 20, 161–170.
  • Moldes, D., Gallego, P., Couto, R. and Sanroman, A., (2003). Grape seeds: the best lignocellulosic waste to produce laccase by solid state cultures of Trametes hirsute. Biotechnology Letters, 25: 491-495.
  • Nicell, J.A., Al-Kassim, L., Bewtra, JK., Taylor, KE.,( 1993). Wastewater treatment by enzyme catalysed polymerization and precipitation. Biodeterior Abstr 7: 1–8.
  • Quan, D., Shin, W. (2004). Amperometric detection of Catechol and Catecholamines by immobilized Laccase from DeniLite. Electroanalysis, 16 (19).
  • Shuttleworth, K. L., ve Bollag, J. M. (1986). Soluble and immobilized laccase as catalysts for the transformation of substituted phenols. Enzym. Microb. Technol. 8, 171–177.
  • Solna´, R., Petr, S.,, (2005). Amperometric flow-Injection determination of phenolic compounds using a biosensor with immobilized Laccase, Peroxidase and Tyrosinase. Electroanalysis 17 (23): 2137–2146.
  • Yamak, O., Kalkan, N.A., Aksoy, S., Altınok, H., Hasırcı, N. (2009). Semi-interpenetrating polymer networks (semi-IPNs) for entrapment of laccase and their use in Acid Orange 52 decolorization. Process Biochem. 44, 440–445.
  • Zawisza, I., Rogalsky, J., Opallo, M., (2006). Electrocatalytic reduction of dioxygen by redox mediator and laccase immobilized in silicate thin film. J Electroanal Chem, 588: 244-252.

Investigation of the optimum conditions of immobilized and free laccase enzyme and its use in decolorization process

Year 2019, Volume: 11 Issue: 1, 153 - 162, 31.01.2019
https://doi.org/10.29137/umagd.508095

Abstract

In this study, Laccase enzyme (L) from Tramatesversicolor was entrapped into poly(N-isopropylacryl amide)-calcium alginate (P(NIPA)-CaAlj)  beads The properties of the immobilized enzyme and use of enzymatic decolorization processes were investigated and compared with those of the free enzyme. Michaelis-Menten constant (Km) and maximum reaction rate (Vmax) values were found to be 1.70x10-2 mM and 2.08x10-3 mM.min-1 for free enzyme respectively. Km and Vmax values were found as 4.80x10-2 mM and 8.70 x 10-3 mM.min-1 for entrapped enzymes respectively. Optimum pH was determined as 5.0 and 6.0 and optimum temperature determined as 40oC and 45oC for free laccase and entrapped laccase respectively. After 30 days of storage at 4 oC free laccase retained 60 % of its original activity. Also after 30 days of storage at 4 oC, entrapped enzymes were retained 83 % its original activity. Immobilized enzyme was used repeatedly 10 times, were retained 77 % of its original activities. Percent decolorization of methyl orange by free enyzme and entrapped enyzmes were found to be 73% and 70%, respectively. 

References

  • Al-Adhami, A.J.H., Bryjak, J., Markiewicz, B.G., Chozch, W.P., (2002). Immobilization of wood-rotting fungi laccases on modified cellulose and acrylic carriers. Process Biochem, 37: 1387-1394.
  • Bamforth, S.M., Singleton, I., (2005). Bioremediation of polycyclic aromatic hydrocarbons: current knowledge and future directions. J Chem Technol Biotechnol 80: 723–736.
  • Curulli A., Cusma A., Kaciulis S., Padeletti G., Pandolfi L., Valentini F., Vitocelli M., (2006). Immobilization of GOD and HRP enzyme on nanostructured substrates. Surf. Interface Anal., 38 478–481.
  • D’Annibale, A., Stazi, A., Vinciguerra, S.R. and Giovannozzi, V.G., (2000). Oxirane-immobilized Lentinula edodes laccase: stability and phenolics removal efficiency in olive mill wastewater. Biotechnol, 77: 265-273.
  • Davis, S., and Burns, R. G. (1990). Decolorization of phenolic effluents by soluble and immobilized phenol oxidases. Appl. Microbiol. Biotechnol. 32, 721–726.
  • Davis, S., and Burns, R. G. (1992). Covalent immobilization of laccase on activated carbon for phenolic effluent treatment. Appl. Microbiol. Biotechnol. 37, 474–479.
  • Fang, H., Huang, J., Ding, L., Li, M., Chen, Z., (2009). Preparation of magnetic Chitosan nanoparticles and immobilization of Laccase. Journal of Wuhan University of Technology-Mater. Sci. Ed.Feb. 24 (1).
  • Freire, R.S., Durán, N., Kubota, L.T., (2001). Effects of fungal laccase immobilization procedures for the development of a biosensor for phenol. Talanta, 54: 681-686.
  • Gianfreda, L., Xu, F., and Bollag, J. M. (1999). Laccases: A Useful Group of Oxidoreductive Enzymes. Bioremediat. J. 3, 1–26.
  • Gokgoz M. and Altinok H., (2012). Immobilization of laccase on polyacrylamide and polyacrylamide - κ - carragennan-based semi-interpenetrating polymer networks. Artificial Cells, Blood Substitutes, and Biotechnology, 40 326–330.
  • Gomez, J., Pazos, M., Couto,R., Sanroman, M., (2005). Chestnut shell and barley bran as potential substrates for lacasse production by Corilopsis rigida under solid-state conditions. Journal of Food Engineering, 68: 315-319.
  • Hu, X., Zhau, X. and Hwang, H., (2007). Comparative study of immobilized Trametes versicolor laccase on nonoparticles and kaolinite. Chemosphere, 66: 1618-1626.
  • Hublik, G. and Schinner, F., (2000). Characterization and immobilization of the Laccase from Pleuratus ostreatus and its use for the continuous elimination of fenolic pollutants. Enzyme Microb Tech, 27: 330-336.,
  • Leonowicz, A., Grzywnowicz, K., (1981). Quantitative estimation of laccase forms in some white-rot fungi using syringaldazine as a substrate. Enzyme Microb Tech, 3: 55-58.
  • Luterek, J., Gianfreda, L., Wojtas-Wasilewska, M., Cho, N., Rogalski, J.,Jaszek, M.,Malarczyk, E.,Staszczak, M.,Fink Boots, M., Leonowicz, A., (1998). Activity of free and immobilized extracellular Cerrena unicolor Laccase in water miscible organic solvents. Cat. Inist, 52: 589-595.
  • Makas Y.G. , Kalkan N.A., Aksoy S., Altinok H., Hasirci N., (2010). Immobilization of laccase in -carrageenan based semi-interpenetrating polymer Networks. Journal of Biotechnology, 148 216–220.
  • Milstein, O., Haars, A., Majerczyck, A., Trojanowski, J., Tautz, D., Zanker, H., Hu¨ ttermann, A. (1988). Removal of Chlorophenols and Chlorolignins from Bleaching Effluent by Combined Chemical and Biological Treatment. Water Sci. Technol. 20, 161–170.
  • Moldes, D., Gallego, P., Couto, R. and Sanroman, A., (2003). Grape seeds: the best lignocellulosic waste to produce laccase by solid state cultures of Trametes hirsute. Biotechnology Letters, 25: 491-495.
  • Nicell, J.A., Al-Kassim, L., Bewtra, JK., Taylor, KE.,( 1993). Wastewater treatment by enzyme catalysed polymerization and precipitation. Biodeterior Abstr 7: 1–8.
  • Quan, D., Shin, W. (2004). Amperometric detection of Catechol and Catecholamines by immobilized Laccase from DeniLite. Electroanalysis, 16 (19).
  • Shuttleworth, K. L., ve Bollag, J. M. (1986). Soluble and immobilized laccase as catalysts for the transformation of substituted phenols. Enzym. Microb. Technol. 8, 171–177.
  • Solna´, R., Petr, S.,, (2005). Amperometric flow-Injection determination of phenolic compounds using a biosensor with immobilized Laccase, Peroxidase and Tyrosinase. Electroanalysis 17 (23): 2137–2146.
  • Yamak, O., Kalkan, N.A., Aksoy, S., Altınok, H., Hasırcı, N. (2009). Semi-interpenetrating polymer networks (semi-IPNs) for entrapment of laccase and their use in Acid Orange 52 decolorization. Process Biochem. 44, 440–445.
  • Zawisza, I., Rogalsky, J., Opallo, M., (2006). Electrocatalytic reduction of dioxygen by redox mediator and laccase immobilized in silicate thin film. J Electroanal Chem, 588: 244-252.
There are 24 citations in total.

Details

Primary Language Turkish
Journal Section Articles
Authors

Haydar Altınok 0000-0002-0043-9385

Publication Date January 31, 2019
Submission Date January 4, 2019
Published in Issue Year 2019 Volume: 11 Issue: 1

Cite

APA Altınok, H. (2019). İmmobilize ve Serbest Lakkaz Enziminin Optimum Koşullarının Araştırılması ve Renk Giderme İşleminde Kullanımı. International Journal of Engineering Research and Development, 11(1), 153-162. https://doi.org/10.29137/umagd.508095

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