Encapsulation of combi-cleas of glycosidases in alginate beads and polyvinyl alcohol for wine aroma enhancement

The aromatic expression of wines can be enhanced by the addition of specific glycosidases, although their poor stability remains a limitation. Coimmobilization of glycosidases as cross-linked enzyme aggregates (combi-CLEAs) offers a simple solution yielding highly stable biocatalysts. Nevertheless, the small particle size of combi-CLEAs hinders their recovery, preventing their industrial application. Encapsulation of combi-CLEAs of glycosidases in alginate beads and in polyvinyl alcohol is proposed as a solution. Combi-CLEAS of β-D-glucosidase and α-L-arabinofuranosidase were prepared and encapsulated. The effects of combi-CLEA loading and particle size on the expressed specific activity (IU/g) of the biocatalysts were evaluated. Best results were obtained with 2.6 mm diameter polyvinyl alcohol particles at a loading of 60 mg/g, exhibiting activities of 1.9 and 1.0 IU/g for β-D-glucosidase and α-L-arabinofuranosidase, respectively. Afterwards, the stability of the biocatalysts was tested in white wine. All the encapsulated biocatalysts retained full activity after 140 incubation days, outperforming both free enzymes and nonencapsulated combi-CLEAs. Nevertheless, the alginate-encapsulated biocatalysts showed a brittle consistency, making recovery unfeasible. Conversely, the polyvinyl-encapsulated biocatalyst remained intact throughout the assay. The encapsulation of combi-CLEAs in polyvinyl alcohol proved to be a simple methodology that allows their recovery and reuse to harness their full catalytic potential

Enzymatic Synthesis of Ascorbyl Palmitate in a Rotating Bed Reactor

Ascorbyl palmitate, an ascorbic acid ester, is an important amphipathic antioxidant that has several applications in foods, pharmaceuticals, and cosmetics. The enzymatic synthesis of ascorbyl palmitate is very attractive, but few efforts have been made to address its process scale-up and implementation. This study aimed at evaluating the enzymatic synthesis of ascorbyl palmitate in a rotating basket reactor operated in sequential batches. Different commercial immobilized lipases were tested, and the most suitable reaction conditions were established. Among those lipases studied were Amano Lipase PS, Lipozyme ® TL IM, Lipozyme ® Novo 40086, Lipozyme ® RM IM and Lipozyme ® 435. Initially, the enzymes were screened based on previously defined synthesis conditions, showing clear differences in behavior. Lipozyme ® 435 proved to be the best catalyst, reaching the highest values of initial reaction rate and yield. Therefore, it was selected for the following studies. Among the solvents assayed, 2-methyl-2-butanol and acetone showed the highest yields, but the operational stability of the catalyst was better in 2-methyl-2-butanol. The tests in a basket reactor showed great potential for large-scale application. Yields remained over 80% after four sequential batches, and the basket allowed for easy catalyst recycling. The results obtained in basket reactor are certainly a contribution to the enzymatic synthesis of ascorbyl palmitate as a competitive alternative to chemical synthesis. This may inspire future cost-effectiveness studies of the process to assess its potential as a viable alternative to be implemented

Enzymatic Synthesis of Ascorbyl Palmitate in a Rotating Bed Reactor

Ascorbyl palmitate, an ascorbic acid ester, is an important amphipathic antioxidant that has several applications in foods, pharmaceuticals, and cosmetics. The enzymatic synthesis of ascorbyl palmitate is very attractive, but few efforts have been made to address its process scale-up and implementation. This study aimed at evaluating the enzymatic synthesis of ascorbyl palmitate in a rotating basket reactor operated in sequential batches. Different commercial immobilized lipases were tested, and the most suitable reaction conditions were established. Among those lipases studied were Amano Lipase PS, Lipozyme® TL IM, Lipozyme® Novo 40086, Lipozyme® RM IM and Lipozyme® 435. Initially, the enzymes were screened based on previously defined synthesis conditions, showing clear differences in behavior. Lipozyme® 435 proved to be the best catalyst, reaching the highest values of initial reaction rate and yield. Therefore, it was selected for the following studies. Among the solvents assayed, 2-methyl-2-butanol and acetone showed the highest yields, but the operational stability of the catalyst was better in 2-methyl-2-butanol. The tests in a basket reactor showed great potential for large-scale application. Yields remained over 80% after four sequential batches, and the basket allowed for easy catalyst recycling. The results obtained in basket reactor are certainly a contribution to the enzymatic synthesis of ascorbyl palmitate as a competitive alternative to chemical synthesis. This may inspire future cost-effectiveness studies of the process to assess its potential as a viable alternative to be implemented

Development of a Hybrid Bioinorganic Nanobiocatalyst: Remarkable Impact of the Immobilization Conditions on Activity and Stability of β-Galactosidase

Hybrid bioinorganic biocatalysts have received much attention due to their simple synthesis, high efficiency, and structural features that favor enzyme activity and stability. The present work introduces a biomineralization strategy for the formation of hybrid nanocrystals from β-galactosidase. The effects of the immobilization conditions were studied, identifying the important effect of metal ions and pH on the immobilization yield and the recovered activity. For a deeper understanding of the biomineralization process, an in silico study was carried out to identify the ion binding sites at the different conditions. The selected β-galactosidase nanocrystals showed high specific activity (35,000 IU/g biocatalyst) and remarkable thermal stability with a half-life 11 times higher than the soluble enzyme. The nanobiocatalyst was successfully tested for the synthesis of galacto-oligosaccharides, achieving an outstanding performance, showing no signs of diffusional limitations. Thus, a new, simple, biocompatible and inexpensive nanobiocatalyst was produced with high enzyme recovery (82%), exhibiting high specific activity and high stability, with promising industrial applications