Enhanced stability and catalytic activity of immobilized α-amylase on modified Fe3O4 nanoparticles

• α-Amylase enzyme was covalently immobilized on the surface of silica-coated modified magnetite nanoparticles.
• Magnetite (Fe3O4) nanoparticles have been considered suitable for immobilization of enzymes.
• The EDX spectrum showed that magnetite nanoparticles were successfully coated with silica.
• The TEM images showed that the diameter of silica-coated magnetite NPs was about 20 nm.

In this work, α-amylase enzyme was covalently immobilized on the surface of silica-coated modified magnetite nanoparticles, for the first time. The synthesis and immobilization process is simple and very fast and consists of the following steps: (1) preparing the magnetic iron oxide nanoparticles using the co-precipitation method, (2) coating NP with silica (SiO2) by sol–gel reaction, (3) preparing the amino-functionalized magnetite NPs by treating silica-coated NPs with 3-aminopropyltriethoxysilane, (4) activating immobilization of α-amylase on the activated amino-functionalized magnetite NPs, and (5) covalently immobilization of α-amylase on the activated-amino-functionalized magnetite NPs. The synthesis steps and characterizations of NPs were examined by FT-IR, XRD, EDX and TEM. The optimum concentration and time for maximum enzyme activity of the immobilized α-amylase are identified to be 150 μg and 4 h, respectively for the hydrolysis of starch. The immobilized α-amylase showed maximal catalytic activity at pH = 6.5 and 45 °C. The kinetic studies shows overall enhancement in the performance of the immobilized enzyme with reference to the free enzyme. Similarly, the thermal stability of the enzyme is found to increase after the immobilization. The Immobilized α-amylase has also been demonstrated to be capable of being reused for six cycles while retaining ∼85% of the initial activity. By using a magnetically active support, quick separation of amylase from reaction mixture is enabled. The Km values were found as 6.27 and 4.77 mM for free and immobilized enzymes, respectively. The Vmax values for the free and immobilized enzymes were calculated as 2.44 and 11.58 μmol/mg min, respectively.