Design of β‐galactosidase/silica biocatalysts: Impact of the enzyme properties and immobilization pathways on their catalytic performance

Abstract

The use of heterogeneous biocatalysis in industrial applications is advantageous and the enzyme stability improvement is a continuous challenge. Therefore, we designed β‐galactosidase heterogeneous biocatalysts by immobilization, involving the support synthesis and enzyme selection (from Bacillus circulans, Kluyveromyces lactis, and Aspergillus oryzae). The underivatized, tailored, macro‐mesoporous silica exhibited high surface area, offered high enzyme immobilization yields and activity. Its chemical activation with glyoxyl groups bound the enzyme covalently, which suppressed lixiviation and conferred higher pH and thermal stability (120‐fold than for the soluble enzyme), without observable reduction of activity/stability due to the presence of silica. The best balance between the immobilization yield (68%), activity (48%), and stability was achieved for Bacillus circulans β‐galactosidase immobilized on glyoxyl‐activated silica, without using stabilizing agents or modifying the enzyme. The enzyme stabilization after immobilization in glyoxyl‐activated silica was similar to that observed in macroporous agarose‐glyoxyl support, with the reported microbiological and mechanical advantages of inorganic supports. The whey lactolysis at pH 6.0 and 25°C by using this catalyst (1 mg ml−1, 290 UI g−1) was still 90%, even after 10 cycles of 10 min, in batch process but it could be also implemented on continuous processes at industrial level with similar results.

Production of High Galacto-oligosaccharides by Pectinex Ultra SP-L: Optimization of Reaction Conditions and Immobilization on Glyoxyl-Functionalized Silica

“…Afterward, the hydrolysis of epoxy groups was carried out by mixing 1 g of solid with 10 mL of 0.1 M H 2 SO 4 for 2 h at 85 °C. The resultant glyceryl-epoxide silica, after being washed with water/acetone (70:30 v/v) and dried, was subjected to a further oxidation step of the glyceryl moieties by reacting with 0.03 M NaIO 4 for 2 h at RT. , In this way, 1 mmol of glyoxyl groups per gram of support was generated, according to the best results reported by Bernal et al The formation of glyoxyl groups on the silica surface was quantified by back-titration with NaHCO 3 /KI, which measured the difference in absorbance at 405 nm of the supernatant before and after the oxidation process …”

In situ immobilization of β‐galactosidase from Bacillus circulans in silica by sol‐gel process: Application in prebiotic synthesis

“…The β–gal B. circulans has been immobilized by adsorption or covalent attachment in organic and inorganic supports . However, each technique has its own advantages and drawbacks, for example the high enzyme lixiviation by the adsorption pathway and the increase of the immobilization costs by covalent attachment due to the synthesis and functionalization of the supports before the immobilization process . Furthermore, in the case the immobilization on organic supports, the biocatalysts have low thermal and mechanic stability, low tolerance towards organic solvents and they are prone to microbial attack .…”

Synthesis of N-Acetyllactosamine and N-Acetyllactosamine-Based Bioactives

“…Silica is an attractive support for enzyme immobilization due to its non-toxicity, microbial resistance, ease of handling and robustness towards high flow rates in continuous processes. [187][188] Engineered enzymes from Trypanosoma rangeli have been immobilized on two silica supports:…”