Use of poly HEMA-MAH -Cu2+ Microbeads for α-Amylase Immobilization
Year 2014,
Volume: 42 Issue: 3, 323 - 330, 01.09.2014
Ayşegül Ayla
Zübeyde Baysal
Abstract
In this article poly 2-hydroxyethyl methacrylate-N-methacryloyl- l -histidin methylester [P HEMA-MAH ] microbeads were prepared by suspension polymerization of 2-hydroxyethyl methacrylate HEMA and N-methacryloyl- l -histidine methyl ester MAH . Some properties of the p HEMA-MAH microbeads were determined by using scanning electron microscopy SEM and swelling tests. Cu2+ ions were chelated on the p HEMA-MAH microbeads, then these microbeads were used in the immobilization of α-amylase in batch system. The maximum α-amylase adsorption capacity of the p HEMA-MAH -Cu2+ beads was found to be 14.88 mg/g at pH 6.0. The optimum temperatures for the immobilized and free enzyme were determined to be 30oC and 25 oC, respectively. Kinetic parameters Km, Vmax of the immobilized enzyme were also determined and compared with those of free enzyme. Reuse studies showed that the immobilized enzyme could reuse six times while retaining 76% of its activity.
References
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- 14. G. Bayramoğlu, M. Yılmaz, M.Y. Arıca, Immobilization of a thermostable α-amylase onto reactive membranes: kinetics characterization and application to continuous starch hydrolysis, Food Chem., 84 (2004) 591.
- 15. A. Kumari, A.M. Kayastha, Immobilization of soybean (Glycine max) α-amylase onto chitosan and amberlite MB-150 beads: optimization and characterization, J. Mol. Catal. B: Enzymatic., 69 (2011) 8.
- 16. M.M. Sarı, C. Armutçu, N. Bereli, L. Uzun, A. Denizli, Monosize microbeads for pseudo-affinity adsorption of human insulin, Colloid. Surface. B., 84 (2011) 140.
- 17. E.B. Altıntas, L. Uzun, A. Denizli, Synthesis and characterization of monosize magnetic poly(glycidyl methacrylate) beads, China Particuol., 5 (2007) 174.
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- 24. A. Derazshamshir, B. Ergün, G. Peşint, M. Odabaşı, Preparation of Zn2+-chelated poly(HEMA-MAH) cryogel for affinity purification of chicken egg lysozyme, J. Appl. Polymer Sci., 109 (2008) 2905.
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- 32. N. Tüzmen, T. Kalburcu, A. Denizli, α-Amylase immobilization onto dye attached magnetic beads: Optimization and characterization, J. Mol. Catal. B: Enzymatic., 78 (2012) 16.
- 33. M.E. Sedaghat, M. Ghiaci, H. Aghaei, S. SoleimanianZad, Enzyme immobilization. Part 3: immobilization of α-amylase on Na-bentonite and modified bentonite, Appl. Clay Sci., 46 (2009) 125.
- 34. J.I. Ida, T. Matsuyama, H. Yamamoto, Immobilization of glucoamylase on ceramic membrane surfaces modified with a new method of treatment utilizing SPCP–CVD, Biochem. Eng. J., 5 (2000) 179.
- 35. I. Gancarz, J. Bryjak, M. Bryjak, G. Poniak, W. Tylus, Plasma modified polymers as a support for enzyme immobilization 1.: allyl alcohol plasma, Eur. Polym. J., 39 (2003) 1615.
- 36. V. Singh, P. Kumar, Carboxymethyl tamarind gum– silica nanohybrids for effective immobilization of amylase, J. Mol. Catal. B: Enzymatic., 70 (2011) 67.
- 37. M.Y. Arica, V. Hasırci, N.G. Alaeddinoğlu, Covalent immobilization of α-amylase onto pHEMA microspheres: preparation and application to fixed bed reactor, Biomaterials., 16 (1995) 761.
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Poli HEMA-MAH -Cu2+ Mikrokürelerin α-Amilaz İmmobilizasyonu için Kullanılması
Year 2014,
Volume: 42 Issue: 3, 323 - 330, 01.09.2014
Ayşegül Ayla
Zübeyde Baysal
Abstract
S Bu çalışmada poli 2-hidroksietil metakrilat-N-metakriloil- L -histidin metilester [P HEMA-MAH ] mikroküreler süspansiyon polimerizasyon yöntemiyle 2-hidroksietil metakrilat HEMA ve N-metakriloil- L -histidin metil ester MAH ile hazırlandı. P HEMA-MAH mikrokürelerin bazı özellikleri taramalı elektron mikroskobu SEM ve şişme testleriyle belirlendi. P HEMA-MAH mikrokürelere Cu2+ bağlandıktan sonra bu mikroküreler α-amilaz immobilizasyonu için kullanıldı. P HEMA-MAH -Cu2+ mikrokürelerin maksimum α-amilaz adsorpsiyon kapasitesi pH 6.0’da 14.88 mg/g olarak bulundu. İmmobilize ve serbest enzim için optimum sıcaklık sırasıyla 30ºC ve 25ºC olarak belirlendi. İmmobilize enzim için kinetik parametreler de Km, Vmax belirlenerek serbest enzim ile karşılaştırıldı. Tekrarlılık çalışmaları, immobilize enzimin orijinal aktivitesinin %76’sını kaybetmeden altı kez tekrar kulanılabilirliğini gösterdi
References
- 1. H. Tümtürk, S. Aksoy, N. Hasırcı, Covalent immobilization of α-amylase onto poly(2-hydroxyethyl methacrylate) and poly(styrene -2-hydroxyethyl methacrylate) microspheres and the effect of Ca2+ ions on the enzyme activity, Food Chem., 68 (2000) 259.
- 2. A. Konieczna-Molenda, A. Kochanowski, A. Walaszek, E. Bortel, P. Tomasik, Immobilization of α-amylase on poly(vinylamine) and poly(vinylformamide) supports and its performance, Chem. Eng. J., 146 (2009) 515.
- 3. S. Sivaramakrishnan, D. Gangadharan, K.M. Nampoothiri, A. Pandey, α-Amylase from microbial sources– an overview on recent developments, Food Technol. Biotechnol., 44 (2006) 173.
- 4. R. Gupta, P. Gigras, H. Mohapatra, V. Kumar Goswami, B. Chauhan, Microbial α-amylases: a biotechnological perspective, Process Biochem., 38 (2003) 1599.
- 5. D. Gangadharan, K. Madhavan Nampoothiri, S. Sivaramakrishnan, A. Pandey, Immobilized bacterial α-amylase for effective hydrolysis of raw and soluble starch, Food Research. Int., 42 (2009) 436.
- 6. A. Pandey, P. Nigam, C.R. Soccol, V.T. Soccol, D. Singh, R. Mohan, Advances in microbial amylases, Biotechnol. Appl. Biochem., 31 (2000) 135.
- 7. J. Bryjak, Glucoamylase, α-amylase and β-amylase immobilisation on acrylic carriers, Biochem. Eng. J., 16 (2003) 347.
- 8. A. Kadziola, J. Abe, B. Svensson, R. Haser, Crystal and molecular structure of barley α-amylase, J. Mol. Biol., 239 (1994) 104.
- 9. P. Tripathi, A. Kumari, P. Rath, A.M. Kayastha, Immobilization of α-amylase from mung beans (Vigna radiata) on Amberlite MB 150 and chitosan beads: A comparative study, J. Mol. Catal. B., 49 (2007) 69.
- 10. M. Safarikova, I. Roy, M.N. Gupta, I. Safarik, Magnetic alginate microparticles for purification of α-amylases, J. Biotechnol., 105 (2003) 255.
- 11. G. Rajagopalan, C. Krishnan, Immobilization of maltooligosaccharide forming α-amylase from Bacillus subtilis KCC103: properties and application in starch hydrolysis, J. Chem. Technol. Biotechnol., 83 (2008) 1511.
- 12. M.E. Çorman, N. Öztürk, N. Bereli, S.Akgöl, A. Denizli, Preparation of nanoparticles which contains histidine for immobilization of Trametes versicolor laccase, J. Mol. Catal. B: Enzymatic., 63 (2010) 102.
- 13. M.V. Kahraman, G. Bayramoğlu, N. KayamanApohan, A. Güngör, α-Amylase immobilization on functionalized glass beads by covalent attachment, Food Chem., 104 (2007) 1385.
- 14. G. Bayramoğlu, M. Yılmaz, M.Y. Arıca, Immobilization of a thermostable α-amylase onto reactive membranes: kinetics characterization and application to continuous starch hydrolysis, Food Chem., 84 (2004) 591.
- 15. A. Kumari, A.M. Kayastha, Immobilization of soybean (Glycine max) α-amylase onto chitosan and amberlite MB-150 beads: optimization and characterization, J. Mol. Catal. B: Enzymatic., 69 (2011) 8.
- 16. M.M. Sarı, C. Armutçu, N. Bereli, L. Uzun, A. Denizli, Monosize microbeads for pseudo-affinity adsorption of human insulin, Colloid. Surface. B., 84 (2011) 140.
- 17. E.B. Altıntas, L. Uzun, A. Denizli, Synthesis and characterization of monosize magnetic poly(glycidyl methacrylate) beads, China Particuol., 5 (2007) 174.
- 18. D. Aktaş Uygun, N. Öztürk, S. Akgöl, A. Denizli, Novel magnetic nanoparticles for the hydrolysis of starch with Bacillus licheniformis α-amylase, J. Appl. Polymer Sci., 123 (2012) 2574.
- 19. A. Kara, B. Osman, H. Yavuz, N. Beşirli, A. Denizli, Immobilization of α-amylase on Cu2+ chelated poly(ethylene glycol dimethacrylate-n-vinyl imidazole) matrix via adsorption, React. Function. Polym., 62 (2005) 61.
- 20. M.A. Abd El-Ghaffar, M.S. Hashem, Immobilization of α-amylase onto chitosan and its amino acid condensation adducts, J. Appl. Polymer Sci., 112 (2009) 805. 21. S. Li, J. Hu, B. Liu, Use of chemically modified PMMA microspheres for enzyme immobilization, Biosystems., 77 (2004) 25.
- 22. G. Bayramoğlu, E. Yalçın, M.Y. Arıca, Immobilization of urease via adsorption onto l-histidine–Ni(II) complexed poly(HEMA-MAH) microspheres: preparation and characterization, Process Biochem., 40 (2005) 3505.
- 23. S. Akgöl, N. Öztürk, A. Denizli, New generation polymeric nanospheres for catalase immobilization, J. Appl. Polymer Sci., 114 (2009) 962.
- 24. A. Derazshamshir, B. Ergün, G. Peşint, M. Odabaşı, Preparation of Zn2+-chelated poly(HEMA-MAH) cryogel for affinity purification of chicken egg lysozyme, J. Appl. Polymer Sci., 109 (2008) 2905.
- 25. N. Ünlü, Ş. Ceylan, M. Erzengin, M. Odabaşı, Investigation of protein adsorption performance of Ni2+-attached diatomite particles embedded in composite monolithic cryogels, J. Sep. Sci., 34 (2011) 2173.
- 26. M. Karataş, S. Akgöl, H. Yavuz, R. Say, A. Denizli, Immunoglobulin G depletion from human serum with metal-chelated beads under magnetic field, Int. J. Biol. Macromol., 40 (2007) 254.
- 27. D. Çimen, A. Denizli, Immobilized metal affinity monolithic cryogels for cytochrome c purification, Colloid. Surface. B., 93 (2012) 29.
- 28. B. Garipcan, A. Denizli, A novel affinity support material for the separation of immunoglobulin G from human plasma, Macromolecular Biosci., 2 (2002) 135.
- 29. M. Yavuz, Z. Baysal, Preparation and use of poly(hydroxyethyl methacrylate) cryogels containing l-histidine for β-casein adsorption, J. Food Sci., 78 (2013) E238.
- 30. P. Bernfeld: α-and β-amylases. In: Methods in Enzymology, S.P. Colowick, N. Kaplan (Eds.), Academic Press, New York, USA (1955) pp. 149-154.
- 31. O.H. Lowry, N.J. Rosebrough, A.L. Farr, R.J. Randall, Protein measurement with the folin phenol reagent, J. Biol. Chem., 193 (1951) 265.
- 32. N. Tüzmen, T. Kalburcu, A. Denizli, α-Amylase immobilization onto dye attached magnetic beads: Optimization and characterization, J. Mol. Catal. B: Enzymatic., 78 (2012) 16.
- 33. M.E. Sedaghat, M. Ghiaci, H. Aghaei, S. SoleimanianZad, Enzyme immobilization. Part 3: immobilization of α-amylase on Na-bentonite and modified bentonite, Appl. Clay Sci., 46 (2009) 125.
- 34. J.I. Ida, T. Matsuyama, H. Yamamoto, Immobilization of glucoamylase on ceramic membrane surfaces modified with a new method of treatment utilizing SPCP–CVD, Biochem. Eng. J., 5 (2000) 179.
- 35. I. Gancarz, J. Bryjak, M. Bryjak, G. Poniak, W. Tylus, Plasma modified polymers as a support for enzyme immobilization 1.: allyl alcohol plasma, Eur. Polym. J., 39 (2003) 1615.
- 36. V. Singh, P. Kumar, Carboxymethyl tamarind gum– silica nanohybrids for effective immobilization of amylase, J. Mol. Catal. B: Enzymatic., 70 (2011) 67.
- 37. M.Y. Arica, V. Hasırci, N.G. Alaeddinoğlu, Covalent immobilization of α-amylase onto pHEMA microspheres: preparation and application to fixed bed reactor, Biomaterials., 16 (1995) 761.
- 38. J.P. Chen, Y.M. Sun, D.H. Chu, Immobilization of α-amylase to a composite temperature-sensitive membrane for starch hydrolysis, Biotechnol. Progg., 14 (1998) 473.
- 39. P.C. Ashly, M.J. Joseph, P.V. Mohanan, Activity of diastase α-amylase immobilized on polyanilines (PANIs), Food Chem., 127 (2011) 1808.