Genipin crosslinked porous chitosan beads as robust supports for β-galactosidase immobilization: Characterization, stability, and bioprocessing potential

ESPARZA-FLORES, ELÍ EMANUEL; CARDOSO, FERNANDA DIAS; SIQUIERA, LARISA BERTOLDO; SANTAGAPITA, PATRICIO R.; HERTZ, PLINHO F.; RODRIGUES, RAFAEL C.

INTERNATIONAL JOURNAL OF BIOLOGICAL MACROMOLECULES

ELSEVIER SCIENCE BV

Año: 2023 vol. 250

0141-8130

This study aimed to modify the porosity of chitosan beads using Na2CO3 as a porogen agent and to crosslink them with genipin for the immobilization of β-galactosidase from Aspergillus oryzae. Immobilization was performed under four different pH conditions (4.5, 6.0, 7.5, and 9.0), resulting in biocatalysts named B4, B6, B7, and B9, respectively. The immobilized enzymes were characterized for immobilization parameters and stability, including thermal, pH, storage, and operational stability. The optimal conditions for the support were determined as 50 mM Na2CO3. The biocatalyst exhibited nearly 100 % retention of initial activity after 5 h of incubation at different pH conditions and showed improved thermal stability compared to the free enzyme across all pH conditions. After 50 cycles of lactose hydrolysis, all biocatalysts retained at least 71 % of their initial activity, with B6 retaining nearly 100 %. Scanning electron microscopy revealed structural modifications, particularly in B4, leading to weakened support structure after reuse. Continuous lactose hydrolysis showed increased productivity from 41.3 to 48.1 g L−1 h−1 for B6, with 78.1 % retention of initial capacity. All biocatalysts retained >95 % activity when stored at 4 °C for 20 weeks, highlighting their suitability for enzyme immobilization in continuous and discontinuous bioprocesses.