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Immobilization of β-glucosidase in fixed bed reactor and evaluation of the enzymatic activity

机译：β-葡萄糖苷酶在固定床反应器中的固定化和酶活性的评价

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摘要

Adsorption of β-glucosidase from almonds, an enzyme with big molecular size (130 kDa, 6.7 nm molecular diameter), on mesoporous SB A-15 silica in fixed bed column was studied. Previously, zeta potential analysis confirmed that the electrostatic interactions between β-glucosidase and SBA-15 were the driving force of the immobilization process. The maximum difference in the zeta potential was 25 mV at pH 3.5. Adsorption isotherm was classified as an L3 (Langmuir type 3) curve according to the Giles classification and fitted to a double Langmuir equation. The adsorbed amount in a fixed bed column was around 3.5 times higher than the amount reached in the adsorption in batch. In addition, the β-glucosidase was strongly immobilized on SBA-15 with only 7 % of leaching in the washing step with buffer solution. Immobilized β-glucosidase was catalytically active in a continuous process, reaching 100 % substrate conversion and maintaining this activity level for more than 10 h without deactivation of the enzyme. Adsorption-desorption isotherms at 77 K before and after the adsorption were carried out, concluding that the adsorption of β-glucosidase was produced blocking the pore mouth, so that a part of the enzyme penetrates inside and another part stays outside the pore.
机译：研究了固定床色谱柱中介孔SB A-15二氧化硅对分子中大分子（130 kDa，分子直径6.7 nm）的β-葡萄糖苷酶的吸附作用。以前，ζ电位分析证实，β-葡萄糖苷酶和SBA-15之间的静电相互作用是固定化过程的驱动力。在pH 3.5下，ζ电位的最大差异为25 mV。吸附等温线根据Giles分类归类为L3（Langmuir类型3）曲线，并拟合到双重Langmuir方程。固定床塔中的吸附量比分批吸附中的吸附量高约3.5倍。此外，β-葡糖苷酶被牢固地固定在SBA-15上，在用缓冲液洗涤的步骤中只有7％的浸出。固定的β-葡萄糖苷酶在连续过程中具有催化活性，可达到100％的底物转化率，并在不使酶失活的情况下保持此活性水平超过10小时。进行吸附前后在77 K的吸附-解吸等温线，得出的结论是，β-葡萄糖苷酶的吸附是阻塞孔口的，一部分酶渗透到内部，另一部分则停留在孔的外部。

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  • 来源
    《Bioprocess and Biosystems Engineering》 |2012年第8期|p.1399-1405|共7页
  • 作者

    J. M. Gomez; M. D. Romero; T. M. Fernandez; E. Diez;

  • 作者单位

    Department of Chemical Engineering,Faculty of Chemistry,Complutense University of Madrid,28040 Madrid, Spain;

    Department of Chemical Engineering,Faculty of Chemistry,Complutense University of Madrid,28040 Madrid, Spain;

    Department of Chemical Engineering,Faculty of Chemistry,Complutense University of Madrid,28040 Madrid, Spain;

    Department of Chemical Engineering,Faculty of Chemistry,Complutense University of Madrid,28040 Madrid, Spain;

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  • 正文语种 eng
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  • 关键词

    SBA-15; β-glucosidase; fixed bed; immobilization; zeta potential;

    机译：SBA-15;β-葡萄糖苷酶;固定床;固定化;ζ电位;

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