Monica Mesa

Application of Hierarchical Porous Silica with a Stable Large Porosity for β‐Galactosidase Immobilization

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.

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

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.

Synthesis of new silicas with high stable and large mesopores and macropores for biocatalysis applications

Monomodal or bimodal porous silicas with large mesopores, constituted by particles or having a monolithic (block type) morphology, respectively, are synthesized using sodium silicate as siliceous species source, cetyltrimethylammonium bromide (CTAB) as pore template and ethyl acetate (EtAc) as pH modifier. The monomodal porosity is represented by 20-30 nm pores and the bimodal one by these pores and also macropores. These characteristics are modulated in function of the CTAB and EtAc concentrations as well as the pH and hydrothermal treatment. The role of these reagents upon the porosity is rationalized. The presence of high CTAB concentration and a rather low pH decreasing rate (function of EtAc concentration and hydrothermal treatment) are essential for having the already known bimodal mesoporous silicas (BMS). On the contrary a rather high pH decreasing rate promotes the formation of the new bimodal mesoporous-macroporous silicas (BMMS) synthesized in this work, where the EtAc also plays the role of emulsion forming agent. The hydrolytic stability of the synthesized silica under aqueous conditions, at different pH values, makes these silicas good candidates for application in different areas of catalysis, especially in the enzymatic one.

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

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.

Synthesis of micron-sized particles of mesoporous silica from tri-block surfactants in presence of fluoride, usable as stationary phase in HPLC

Abstract The relations between the morphology (particle shape and size) of SBA-15 and SBA-16 mesoporous silica and the synthesis parameters (composition of the reaction mixtures, nature of the triblock surfactant, fluoride anions, synthesis procedure) were established. This allowed defining the conditions for the formation of materials with micro-sized isometric particles suitable for HPLC applications. The presence of fluoride anions favors the formation of spherical shaped particles, with probably higher weak acidic silanol groups content to give higher hydrophobic materials after silylation, which is an advantage for their use as HPLC reverse phases. The different materials were evaluated in HPLC tests (separation of mixtures of aromatic and polynuclear aromatic molecules).

Formation mechanism of SBA-3, SBA-15 and SBA-16 type mesoporous silica in acidic solutions

Abstract The evolution of the species in acidic solutions, prepared from tetraethoxysilane (TEOS), surfactant (Cetyltrimethylammonium bromide, Pluronic P123 or Pluronic F127) and HCl, was followed by visual observation and Dynamic Light Scattering (DLS) measurements between 25 and 90°C until their precipitation. The nature of the precipitated phases was studied by optical microscopy (in-situ in the reaction mixtures) and by SEM (after filtration and drying). In all cases, the first stage corresponds to the formation of micelles coated by siliceous species (composite colloids). In a second stage the modification of their colloidal characteristics leads to their aggregation with a phase separation into micron-sized liquid particles. In the third stage, the progress of the polycondensation transforms the latter into solid particles of mesoporous silica. The final morphology depends on the polycondensing degree of the silica before the phase separation and on the prevailing synthesis parameters (temperature and pH).

Catalytic conversion of α-pinene by using mesoporous aluminosilicates

Mesoporous aluminosilicates are prepared using CTMABr and sodium silicate and aluminum sulfate as Si and Al source, respectively. The materials are characterized by XRD, elemental analysis, N 2 adsorption, 27 Al NMR and Infrared spectroscopy with pyridine sorption. The results show that the use of aluminum sulfate conducts to mesoporous aluminosilicates where the framework charges are compensated by protons giving acidic Bronsted sites No Lewis acid sites are observed. The Bronsted acidity seems not to be directly related to the number of incorporated tetrahedral Al atoms but to the nature of the Bronsted sites and to the mesoporous organization. Their catalytic activity towards the transformation of α-pinene is due to their Bronsted acidity. A good selectivity at 160°C towards camphene (75%) can be obtained with the catalyst with the lowest Si/Al ratio.

Use of mesoporous silica as a reinforcing agent in rubber compounds

Abstract Mesoporous silica is used as filler for styrene-butadiene rubber (SBR); filler-polymer interactions are compared with those exhibited when Ultrasil silica (VN3) is used. A silane coupling agent is added to improve filler dispersion and its influence on the bound-rubber formation is also investigated. The bound-rubber content is higher for the mesoporous silica and increases further for the sample containing silane. The increase is explained by chemical interactions between filler and rubber and penetration of the rubber chains into the mesopores. This is confirmed by 13C solid-state NMR, IR spectroscopy and differential scanning calorimetry. Dynamic mechanical thermal analysis shows higher storage modulus for the rubber filled with mesoporous silica.

Effect of the porosity and hydrothermal stability of SBA-16 type mesoporous silica on the characteristics of their carbon replicas

The use of hard templates is one of the best ways for obtaining ordered mesoporous carbons (OMC) with well controlled morphology and porosity, characteristics that are very important for their use in many technological applications. Since the SBA-16 type mesoporous silica has a 3D connected mesostructure and can be synthesized with different morphologies, we select this material as a hard template and study the influence of its pore (cavities and entrances) size and hydrothermal stability on the qualities of the OMC replicas obtained by liquid impregnation of sucrose.

Adaptation of silanol groups in mesoporous silica to be silylated and their evaluation by ftir

Abstract Procedures reported in the literature to modify the silanol content of mesoporous silica are applied to a material prepared with the triblock copolymer Pluronic F127 in acidic medium under quiescent conditions, in order to adapt it for silylation reaction. These include: calcination at relatively high temperature and rehydroxylation in presence of NH 4 OH. The nature and content of silanol groups in these materials is measured by following the pyridine interaction with them through the infrared specific absorptions at 1595 and 1446 cm −1 assigned to pyridine interacting with hydrogen-bonded and free silanol groups respectively. In order to render these SBA-16-type mesoporous silica hydrophobic, a silylation reaction is effected in which its efficiency depends mainly on the nature and concentration of silanol groups. To follow the degree of silylation a method based on infrared transmission spectroscopy (FTIR) is designed in this work. With this method, unlike solid state 29 Si-NMR, it is possible to measure the relative concentrations of the organic groups bonded to silica when two silylating reagents are used, and to qualify in an indirect way how the weak silanol groups change during the different post synthesis treatments of a mesoporous silica.