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

• Published in: International journal of biological macromolecules 2023-10, Vol.250, p.126234-126234, Article 126234
• Main Authors: Esparza-Flores, Elí Emanuel, Cardoso, Fernanda Dias, Siquiera, Larisa Bertoldo, Santagapita, Patricio R., Hertz, Plinho F., Rodrigues, Rafael C.
• Format: Article
• Language: English
• Subjects: Batch reactor; Chitosan; Column reactor; Enzyme immobilization; Genipin; β-Galactosidase
• ISSN: 0141-8130; 1879-0003
• DOI: 10.1016/j.ijbiomac.2023.126234

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. •Addition of Na2CO3 modified the porosity of chitosan beads and its superficial area.•The biocatalysts performed better thermal stability than the free enzyme.•The structure of the biocatalysts was modified according to the pH.•Biocatalysts retained at least 71 % of their activity after 50 cycles for lactose hydrolysis.•Biocatalysts presented great stability during the continuous bioprocess for 20 days.