Magnetic Polymeric Hybrid Microspheres as Adsorbent Supports for Laccase Immobilization

Magnetic polymeric hybrid microspheres based on poly(vinylphenylboronic acid-co-divinylbenzene), Fe3O4, and TiO2 [P(VBA-co-DVB)/Fe3O4-TiO2] were synthesized and investigated as adsorbent supports for laccase immobilization. The physicochemical, morphological, structural, and surface characteristics of the microspheres were evaluated by SEM, FT-IR, BET, XRD, and zeta potential analyses. The resulting polymeric hybrid microspheres exhibited a porous polymeric architecture, magnetic responsiveness, and functional surface groups capable of promoting enzyme–support interactions. A BET surface area of 50.26 m2 g−1 was obtained, with BJH average pore diameters of 16.53 and 36.11 nm for the adsorption and desorption branches, respectively. Laccase was directly immobilized from an ultrafiltered crude enzyme preparation without additional chromatographic purification or the use of an external cross-linking agent, predominantly through adsorption and multipoint non-covalent interactions. Approximately 79% of the initial soluble laccase activity was removed from the liquid phase during immobilization. The catalytic properties of free and immobilized laccase were comparatively evaluated in terms of pH-dependent activity, apparent kinetic parameters, thermal stability, storage stability, and operational reusability. Immobilization altered the apparent kinetic response of the enzyme, improved activity retention during storage, and provided partial, temperature-dependent protection against thermal inactivation. In addition, the magnetic nature of the polymeric hybrid microspheres enabled rapid recovery and repeated use of the immobilized biocatalyst, which retained 43% of its initial activity after 20 catalytic cycles. These findings demonstrate that the developed magnetic polymeric hybrid microspheres provide a recoverable and reusable platform for direct laccase immobilization from ultrafiltered crude preparations and may offer advantages for simplified enzyme processing, reusable biocatalysis, and environmental applications.