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The enzyme encapsulation is a very well‐known stabilization pathway. However, there are some challenges in order to avoid the enzyme denaturation under encapsulation conditions. The β‐galactosidase from Bacillus circulans was immobilized through sol‐gel encapsulation route assisted by Triton X‐100 surfactant and sugars. The effects of sugar presence in the immobilization process and the gelation time on the biocatalyst activity/stability were explained taking into account the characteristics of the formed silica matrix and the changes of the enzyme environment. The enzyme was effectively immobilized by this strategy, with high immobilization yield in terms of activity (29%) and expressed activity (47 IU/g). The immobilization through silica sol‐gel in the presence of 1×10−3 M Triton X‐100 and fructose conferred 28.4‐fold higher stability to the enzyme compared with the soluble form. This is an advantage for its use in the synthesis of the galacto‐oligosaccharides at 50ºC. The total lactose conversion to galacto‐oligosaccharides was 26%wt, which is comparable with that reported in the literature. The obtained biocatalyst is useful for the synthesis of galacto‐oligosaccharides and its catalytic behavior is rationalized in this work.

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.

Hierarchical porous silica, particles or monoliths, were synthesized by polycondensation of sodium silicate in the presence of cetyltrimethylammonium bromide and ethyl acetate at different concentrations under hydrothermal conditions. They were used as the support for the immobilization of β-galactosidase from Kluyveromyces lactis by adsorption. The enzyme loading capacity (higher than 50 mg g−1 support) and the retention ability (lixiviation less than 20 % after 72 h of catalysis) of these supports were explained as a function of the hierarchical porosity, mesopore sizes of 10–40 nm, and macropore sizes of 0.07–20 μm and the presence of ionized silanol groups on the surface. The optimum pH value and temperature for the maximum activity of the obtained hybrid biocatalyst were evaluated, indicating that the three-dimensional structure of the lactase was not significantly affected during the immobilization process. The stability under extreme conditions was improved in comparison with soluble lactase. The porous supports exhibited morphological and porous stability under the immobilization and catalytic processes. These results show that the obtained materials are good candidates for the immobilization of large enzymes, such as β-galactosidase.

The mesoporous silica have been considered fascinating materials for many techonological applications due to their porous and morphological characteristics. This review focuses on their use as stationary phases for liquid chromatography, supports for immobilizing biomolecules, catalysts, agent for polymer reinforcement and hard templates for the preparation of mesoporous carbons.

Chitosan was derivatized by two methodologies for analyzing their effect on chitosan physicochemical characteristics and its applicability as carrier for Bacillus circulans b-galactosidase immobilization. Glutaraldehyde (GA) and epichlorohydrin (EPI) were used for crosslinking and activation of chitosan, producing the corresponding supports (C-GA and C-EPI-EPI) after a one-step and a two- step process, respectively. The spherical shape and mean diameter of chitosan particles was not significantly affected by polymer derivati- zation, while Fourier transform infrared analysis showed that in both cases, chitosan polymer was chemically modified. TGA analysis indicated that C-EPI-EPI was the most thermally stable. The high degree of activation of C-EPI-EPI (586 lmol of aldehydes/g) resulted in the highest loss of activity during immobilization; hence a support with 100 lmol of aldehydes/g was produced (C-EPI-EPI100). The highest expressed activity (89.3 IU/g) was obtained with the enzyme immobilized in C-GA, while the biocatalyst with highest thermal stability at 60 � C was obtained with C-EPI-EPI100 (half-life was 84-fold higher than the one of the soluble enzyme). The best compro- mise between biocatalyst expressed activity and thermal stability corresponded to b-galactosidase immobilized in C-EPI-EPI100. Accord- ing to this study, chitosan derivatized with EPI is a thermally stable carrier appropriate for producing highly stable immobilized B. circulans b-galactosidase. V C 2013 Wiley Periodicals, Inc. J. Appl. Polym. Sci. 2014, 131, 40171.