Encapsulation, immobilization

Self-organized materials with high surface area and pore size 3-25 nm was produced used templating and coassembly. The highly porous nature of the ordered combined with low adsorption and emission in the visible spectrum, facile diffusion makes them good candidate for optical and chemical sensor and provide new avenues for encapsulation/ immobilization processes and solve the problems mentioned above. 

Encapsulation. Immobilization of enzymes by encapsulation within semipermeable structures dates back to the 1970s. There are three fundamental variations of this approach. In coacervation, aqueous microdroplets containing the enzyme are suspended in a water-immiscible solvent containing a polymer, such as cellulose nitrate, polyvinylacetate, or polyethylene. A solid film of polymer can be induced to form at the interface between the two phases, thereby producing a microcapsule containing the enzyme. A second approach involves interfacial polymerization in which an aqueous solution of the enzyme and a monomer are dispersed in an immiscible solvent with the aid of a surfactant. A second (hydrophobic) monomer is then added to the solvent and condensation polymerization is allowed to proceed. This approach has been used extensively with nylons, but is also applicable to polyurethanes, other polyesters, and polyureas. 

Yilmaz, E., and M. Sezgin. 2012. Enhancement of the Activity and Enantioselectivity of Lipase by Sol-Gel Encapsulation Immobilization onto B-Cyclodextrin-Based Polymer. Applied Biochemistry and Biotechnology 166 (8) 1927-1940. 

The resolution of racemic ethyl 2-chloropropionate with aliphatic and aromatic amines using Candida cylindracea lipase (CCL) [28] was one of the first examples that showed the possibilities of this kind of processes for the resolution of racemic esters or the preparation of chiral amides in benign conditions. Normally, in these enzymatic aminolysis reactions the enzyme is selective toward the (S)-isomer of the ester. Recently, the resolution ofthis ester has been carried out through a dynamic kinetic resolution (DKR) via aminolysis catalyzed by encapsulated CCL in the presence of triphenylphosphonium chloride immobilized on Merrifield resin (Scheme 7.13). This process has allowed the preparation of (S)-amides with high isolated yields and good enantiomeric excesses [29]. 

Very few examples have been described for the non-covalent immobilization of chiral porphyrin complexes (Fig. 26). In the first case, the porphyrin-dichlororutheninm complex was encapsulated in silica, which was prepared around the complex by a sol-gel method [78], in an attempt to prevent deactivation observed in solution in the epoxidation of different alkenes with 2,6-dichloropyridine N-oxide. In fact, the heterogeneous catalyst is much more active, with TON up to 10 800 in the case of styrene compared to a maximum of 2190 in solution. Enantioselectivities were about the same imder both sets of conditions, with values aroimd 70% ee. 

Dynamic light-scattering experiments or the analysis of some physicochemical properties have shown that finite amounts of formamide, A-methylformamide, AA-dimethyl-formamide, ethylene glycol, glycerol, acetonitrile, methanol, and 1,2 propanediol can be entrapped within the micellar core of AOT-reversed micelles [33-36], The encapsulation of formamide and A-methylformamide nanoclusters in AOT-reversed micelles involves a significant breakage of the H-bond network characterizing their structure in the pure state. Moreover, from solvation dynamics measurements it was deduced that the intramicellar formamide is nearly completely immobilized [34,35],... 

Encapsulation of [Co(bpy)3]2+ within zeolite frameworks has also been shown to have a remarkable influence on the electronic spin state of the complex.240 Distortions imparted on the tris-chelate complex by the confines of the zeolite supercage are found to be responsible for stabilizing the unusual low-spin electronic ground state.241,242 The [Co(bpy)3]3+/2+ couple has been measured for the encapsulated complex and it has been found that the complexes remain within the zeolite and do not exchange with the bulk solution.243 Electrochemistry of [Co(bpy)3]3+/2+ immobilized within a sol-gel has also been studied.244... 

Immobilization of chiral complexes in PDMS membranes offers a method for the generation of new chiral catalytic membranes. The heterogenization of the Jacobsen catalyst is difficult because the catalyst loses its enantioselectivity during immobilization on silica or carbon surfaces whereas the encapsulation in zeolites needs large cages. However, the occlusion of this complex in a PDMS matrix was successful.212 The complex is held sterically within the PDMS chains. The Jacobsen catalyst occluded in the membrane has activity and selectivity for the epoxidation of alkenes similar to that of the homogeneous one, but the immobilized catalyst is recyclable and stable. 

In a similar vein, Gelman et al. [25] demonstrated encapsulation by the use of two SiC>2 sol-gel matrices [26,27] to immobilize two catalysts Wilkinson s catalyst and an amine that acts as a base catalyst. The reaction is illustrated in Scheme 5.10. 

Figure 5.1 The use of multiple catalysts encapsulated in star polymers to enable a one-pot cascade reaction containing interfering catalysts. Catalyst 11 is formed via the protonation of imidazolidinone (9) by immobilized PSTA (10), and is responsible for the nucleophilicaddition ofN-methylindole (14)...

Common to all encapsulation methods is the provision for the passage of reagents and products through or past the walls of the compartment. In zeolites and mesoporous materials, this is enabled by their open porous structure. It is not surprising, then, that porous silica has been used as a material for encapsulation processes, which has already been seen in LbL methods [43], Moreover, ship-in-a-bottle approaches have been well documented, whereby the encapsulation of individual molecules, molecular clusters, and small metal particles is achieved within zeolites [67]. There is a wealth of literature on the immobilization of catalysts on silica or other inorganic materials [68-72], but this is beyond the scope of this chapter. However, these methods potentially provide another method to avoid a situation where one catalyst interferes with another, or to allow the use of a catalyst in a system limited by the reaction conditions. For example, the increased stability of a catalyst may allow a reaction to run at a desired higher temperature, or allow for the use of an otherwise insoluble catalyst [73]. 

Enzymatic degradation was tested with commercial LAC from M. thermophila (2,000 U L ). E2 and EE2 were completely degraded even in the absence of mediators after 3 and 5 h, respectively, and after 1 h in the presence of some mediators. For El total removal was achieved in 8 h in the presence of VA and >70% for the other mediators after 24 h, whereas elimination reached 65% in the absence of mediators [8]. The immobilization of this enzyme by encapsulation in a sol-gel matrix [58] was employed for the treatment of a mixture of El, E2, and EE2 both in a batch stirred tank reactor (BSTR) operating in cycles and a continuous PBR. Removal of estrogens was >85% in the BSTR and 55%, 75%, and 60% for El, E2, and EE2, respectively, in the PBR. Both systems were able to reduce the estrogenic activity of the mixture in 63%. Likewise, the immobilization of VP in the form of CLEAs completely removed E2 and EE2 within 10 min from batch experiments, with a concomitant reduction of estrogenic activity, higher than 60% for both compounds [44]. 

The first belief in the possibility of enzyme stabilization on a silica matrix was stated by Dickey in 1955, but he did not give experimental evidence, only mentioning that his experiments were unsuccessful [65]. A sol-gel procedure for enzyme immobilization in silica was first developed by Johnson and Whateley in 1971 [66]. The entrapped trypsin retained about 34 % of its tryptic activity observed in solution before the encapsulation. Furthermore, the enzyme was not released from the silica matrix by washing, demonstrating the increased stability and working pH range. Unfortunately, the article did not attract attention, although their method contained all the details that may be found in the present-day common approach. This was probably due to its publication in a colloid journal that was not read by biochemists. 

The main advantage is that the entrapment conditions are dictated by the entrapped enzymes, but not the process. This includes such important denaturing factors as the solution pH, the temperature and the organic solvent released in the course of precursor hydrolysis. The immobilization by THEOS is performed at a pH and temperature that are optimal for encapsulated biomaterial [55,56]. The jellification processes are accomplished by the separation of ethylene glycol that possesses improved biocompatibility in comparison with alcohols. 

Wei, Y., Xu, J., Feng, Q., Lin, M., Dong, H., Zhang, W.-J. and Wang, C. (2001) A novel method for enzyme immobilization direct encapsulation of add phosphatase in nanoporous silica host materials. Journal of Nanoscience and Nanotechnology, 1, 83-93. 

One of the most promising applications of enzyme-immobilized mesoporous materials is as microscopic reactors. Galameau et al. investigated the effect of mesoporous silica structures and their surface natures on the activity of immobilized lipases [199]. Too hydrophilic (pure silica) or too hydrophobic (butyl-grafted silica) supports are not appropriate for the development of high activity for lipases. An adequate hydrophobic/hydrophilic balance of the support, such as a supported-micelle, provides the best route to enhance lipase activity. They also encapsulated the lipases in sponge mesoporous silicates, a new procedure based on the addition of a mixture of lecithin and amines to a sol-gel synthesis to provide pore-size control. 

This encapsulation procedure gives the highest activity for the lipases. The lecithin/ amines mixture structuring the pore network leads to a suitable phospholipids bilayer-like environment, which avoids the necessity to create an interface by substrate assembly. Monduzzi and coworkers compared the activity of lipase that was immobilized on SBA-15 physically, or chemically with glutardialdehyde [200]. 

Recently, we proposed an alternative process for encapsulating biomacromolecules within PE microcapsules. This approach involves using nanoporous particles as sacrificial templates for both enzyme immobilization and PE multilayer capsule formation (Figure 7.2, route (I)) [66,67]. Unlike previous LbL encapsulation strategies, this approach is not limited to species that undergo crystallization, and is not dependent upon adjustments in electrostatic interactions within PE microcapsules to alter shell permeability characteristics. The salient feature of this method is that it is applicable to a wide range of materials for encapsulation. 

In general, biomolecules such as proteins and enzymes display sophisticated recognition abilities but their commercial viability is often hampered by their fragile structure and lack of long term stability under processing conditions [69]. These problems can be partially overcome by immobilization of the biomolecules on various supports, which provide enhanced stability, repetitive and continuous use, potential modulation of catalytic properties, and prevention of microbial contaminations. Sol-gel and synthetic polymer-based routes for biomolecule encapsulation have been studied extensively and are now well established [70-72]. Current research is also concerned with improving the stability of the immobilized biomolecules, notably enzymes, to increase the scope for exploitation in various... 

In order to overcome some limitations of the adsorption process due to surface accessibility or diffusional hindering, immobilization of enzymes by direct in situ encapsulation has been investigated. When inorganic supports can be prepared in mild conditions compatible with the enzyme stability, then such processes allow... 

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