Synthesis of organic-inorganic hybrid nanoflowers of lipases from Candida antarctica type B (CALB) and Thermomyces lanuginosus (TLL): Improvement of thermal stability and reusability

Enzyme immobilization is used to improve the application of enzymes, allowing the reuse of biocatalysts and increasing their stability under reaction conditions. Immobilization of enzymes through structures, such as nanoflowers, is an innovative, simple, and low-cost method compared to other techniques. In this context, the main objective of this work is to synthesize hybrid biocatalytic nanostructures, similar to flowers, of lipases from Candida antarctica type B (CALB) and Thermomyces lanuginosus (TLL). The production of nanoflowers occurred by precipitation of lipases with CuCl2 or CuSO4 salts for 72 h. However, challenges and obstacles were faced in obtaining effective and practical nanoflowers, such as nanoflowers' low thermal stability and reusability. To overcome these challenges, two conditions were tested: nanoflowers cross-linked with glutaraldehyde and nanoflowers and nanoparticles cross-linked with glutaraldehyde. This last biocatalyst prepared by CuSO4 precipitation showed better thermal stability (half-life about 230 and 233 min for CALB and TLL, respectively, under incubation at 60 °C and pH 7). The CALB biocatalyst retained 70 % of its initial activity (2.31 U) after 10 cycles of hydrolysis. Therefore, this work shows not only the problems and barriers of nanoflowers synthesis, but also the possibility of producing more stable and efficient biocatalysts using improved protocols.