Carboxyl-functionalized magnetic nanoparticles were synthesized via chemical co-precipitation
method and modified with oleic acid which was oxidized by potassium permanganate, and κ-carrageenase
sp. ASY5 was subsequently immobilized onto them. The immobilization conditions
were further optimized, and the characterizations of the immobilized κ-carrageenase were investigated.
The κ-carrageenase was immobilized onto magnetic iron oxide nanoparticles, and the bonding
was verified by Fourier transform infrared spectroscopy. The optimal conditions for κ-carrageenase
immobilization were 2.5% (w/v) glutaraldehyde, 13.9 U κ-carrageenase for 20 mg of magnetic nanoparticles,
a 2-h cross-linking time, and a 2-h immobilization time at 25°C. Under these conditions, the activity of the
immobilized enzyme and the enzyme recovery rate were 326.0 U · g-1
carriers and 46.9%, respectively. The
properties of the immobilized κ-carrageenase were compared with those of the free enzyme. The optimum
temperatures of the free and immobilized κ-carrageenase were 60 and 55°C, respectively, and the optimum
pH of κ-carrageenase did not change before and after immobilization (pH 7.5). After immobilization,
κ-carrageenase exhibited lower thermal stability and improved pH stability, as well as better storage stability.
The immobilized κ-carrageenase maintained 43.5% of the original activity after being used 4 times. The
kinetic constant value (Km) of κ-carrageenase indicates that the immobilized enzyme had a lower binding
affinity for the substrate.
Under optimal conditions, the activity of the immobilized enzyme and enzyme recovery rate were
326.0 U · g-1
·κ-carrageenase-CMNPs and 46.9%, respectively. The thermal, pH, and storage stabilities of
κ-carrageenase-CMNPs were relatively higher than those of free κ-carrageenase.