Reversible Immobilization of Catalase by Metal Chelate Affinity Interaction on Magnetic Beads
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- 作者：Mü ; frettin Sari ; Sinan Akgö ; l ; Melike Karata
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- 刊名：Industrial & Engineering Chemistry Research
- 出版年：2006
- 出版时间：April 26, 2006
- 年：2006
- 卷：45
- 期：9
- 页码：3036 - 3043
- 全文大小：179K
- 年卷期：v.45,no.9(April 26, 2006)
- ISSN：1520-5045
文摘
A magnetic metal-chelate adsorbent utilizing N-methacryloyl-(L)-cysteine methyl ester (MAC) as a metal-chelating ligand was prepared. MAC was synthesized using methacryloyl chloride and L-cysteine methylester dihydrochloride. Magnetic beads with an average size of 150-250 
m were obtained by suspensionpolymerization of 2-hydroxyethyl methacrylate (HEMA) and MAC carried out in a dispersion medium. Mag-poly(HEMA-MAC) beads were characterized by surface area measurements, swelling tests, electron spinresonance (ESR) spectroscopy, elemental analysis, and scanning electron microscopy (SEM). The specificsurface area of the magnetic beads was found to be 92.6 m2/g. Elemental analysis of MAC for nitrogen wasestimated as 55.4 mol/g. Then, Fe3+ ions were chelated on the magnetic beads. The Fe3+ loading was 12.7mol/g of support. Fe3+-chelated magnetic beads with a swelling ratio of 62% were used in the immobilizationof catalase in a batch system. This approach to the preparation of enzyme carrier has several advantages overconventional immobilization methods. An expensive, time-consuming, and critical step in the preparation ofimmobilized metal-affinity carriers is the coupling of a chelating ligand to the adsorption matrix. In thisprocedure, comonomer MAC acts as the metal-chelating ligand, and it is possible to load metal ions directlyon the beads without further modification steps. The maximum catalase immobilization capacity of the mag-poly(HEMA-MAC)-Fe3+ beads was observed to be 192 mg/g at pH 5.5. The Km value for immobilizedcatalase [mag-poly(HEMA-MAC)-Fe3+] (32.6 mM) was higher than that for free catalase (22.8 mM).Immobilized catalase exhibits enhanced stability in reaction conditions over a wide pH range (pH 5.5-7.0)and retains an activity of 76% after 10 successive batch reactions, demonstrating the usefulness of the enzyme-loaded magnetic beads in biocatalytic applications. The optimum temperature for the immobilized preparationof mag-poly(HEMA-MAC)-Fe3+-catalase at 45 
C was 5 C higher than that of the free enzyme at 40 C.Storage stability was found to increase with immobilization.
m were obtained by suspensionpolymerization of 2-hydroxyethyl methacrylate (HEMA) and MAC carried out in a dispersion medium. Mag-poly(HEMA-MAC) beads were characterized by surface area measurements, swelling tests, electron spinresonance (ESR) spectroscopy, elemental analysis, and scanning electron microscopy (SEM). The specificsurface area of the magnetic beads was found to be 92.6 m2/g. Elemental analysis of MAC for nitrogen wasestimated as 55.4 mol/g. Then, Fe3+ ions were chelated on the magnetic beads. The Fe3+ loading was 12.7mol/g of support. Fe3+-chelated magnetic beads with a swelling ratio of 62% were used in the immobilizationof catalase in a batch system. This approach to the preparation of enzyme carrier has several advantages overconventional immobilization methods. An expensive, time-consuming, and critical step in the preparation ofimmobilized metal-affinity carriers is the coupling of a chelating ligand to the adsorption matrix. In thisprocedure, comonomer MAC acts as the metal-chelating ligand, and it is possible to load metal ions directlyon the beads without further modification steps. The maximum catalase immobilization capacity of the mag-poly(HEMA-MAC)-Fe3+ beads was observed to be 192 mg/g at pH 5.5. The Km value for immobilizedcatalase [mag-poly(HEMA-MAC)-Fe3+] (32.6 mM) was higher than that for free catalase (22.8 mM).Immobilized catalase exhibits enhanced stability in reaction conditions over a wide pH range (pH 5.5-7.0)and retains an activity of 76% after 10 successive batch reactions, demonstrating the usefulness of the enzyme-loaded magnetic beads in biocatalytic applications. The optimum temperature for the immobilized preparationof mag-poly(HEMA-MAC)-Fe3+-catalase at 45 C was 5 C higher than that of the free enzyme at 40 C.Storage stability was found to increase with immobilization.