Improved biocatalysts based on Bacillus circulans β-galactosidase immobilized onto epoxy-activated acrylic supports: Applications in whey processing

β-Galactosidase from Bacillus circulans was immobilized onto Eupergit C, C 250 L, and Sepabeads EP, at low (3 mg/g of gel), medium (30 mg/g of gel) and high applied loads (120 mg/g of gel). Comparison of the best conditions for the immobilization process at 25 °C showed that Eupergit C and C 250 L require milder conditions than Sepabeads EP (e.g. lower ionic strength and pH, shorter incubation time) to achieve immobilized activity yields over 90%.Seeking to increase the thermostability of the derivatives, the effects of two post-immobilization treatments were studied: alkaline incubation to favor the formation of additional covalent linkages, and blocking of excess oxirane groups by reacting with β-mercaptoethanol or glycine. The blocking process improved the thermal stability of all derivatives, with glycine having a greater effect than β-mercaptoethanol. Thermal stability further improved when an alkaline incubation was performed before blocking. The stabilization achieved by this combined strategy was more effective for Eupergit-based than for Sepabeads EP derivatives, and was higher at medium applied load (30 mg/g of gel). Thus, stabilized biocatalysts obtained at medium load on Eupergit C and C 250 L showed half-lives 10-fold higher than similar Sepabeads derivatives (387, 385 and 37 h, respectively).These improved biocatalysts were used batchwise and in continuous processes for lactose hydrolysis in cheese wheys. Similar degrees of lactose hydrolysis were achieved with both Eupergit derivatives, but they varied depending on the substrate, being highest for Mozzarella cheese whey. The degree of lactolysis achieved at 50 °C was between 80 and 90% after 2 h, and derivatives could be reused batchwise for at least 6 cycles.

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► β-Galactosidase from Bacillus circulans was immobilized onto Eupergit C and Sepabeads EP.
► Thermostability was improved by alkaline treatment and blocking with glycine.
► Eupergit-based derivatives showed half-life 10-fold higher than Sepabeads derivatives.
► Improved derivatives were used for lactose hydrolysis in cheese whey at 50 °C.
► The enzymatic derivatives could be reused batchwise for at least 6 cycles.