Soluble and Cross-Linked Aggregated Forms of α-Galactosidase from Vigna mungo Immobilized on Magnetic Nanocomposites: Improved Stability and Reusability.

α-Galactosidases hold immense potential due to their biotechnological applications in
various industrial and functional food sectors. In the present study, soluble and covalently
cross-linked aggregated forms of a low molecular weight, thermo-labile α-galactosidase
from Vigna mungo (VM-αGal) seeds were immobilized onto chitosan-coated magnetic
nanoparticles for improved stability and repeated usage by magnetic separation. Parameters like precipitants (type, amount, and ratio), glutaraldehyde concentration, and enzyme
load were optimized for the preparation of chitosan-coated magnetic nanocomposites of
cross-linked VM-αGal (VM-αGal-MC) and VM-αGal (VM-αGal-M) resulted in 100%
immobilization efficiency. Size and morphology of VM-αGal-M were studied through
dynamic light scattering (DLS) and scanning electron microscopy (SEM), while Fourier
transform infrared spectroscopy (FTIR) was used to study the chemical composition of
VM-αGal-MC and VM-αGal-M. VM-αGal-MC and VM-αGal-M were found more
active in a broad range of pH (3–8) and displayed optimal temperatures up to 25 °C higher
than VM-αGal. Addition of non-ionic detergents (except Tween-40) improved VM-αGalMC activity by up to 44% but negatively affected VM-αGal-M activity. Both VM-αGalMC (15% residual activity after 21 min at 85 °C, Ed 92.42 kcal/mol) and VM-αGal-M
(69.0% residual activity after 10 min at 75 °C, Ed 39.87 kcal/mol) showed remarkable
thermal stability and repeatedly hydrolyzed the substrate for 10 cycles.