Entrapment and Encapsulation

Entrapment can be defined as any system in which an enzyme or whole cell is physically restricted within a confined space or network. This class of immobilization is often extended to include systems where a combination of physical entrap- 

Property Examples or typical range Characteristics and considerations 

Binding mode Covalent, ionic or physical adsorption. Pre-activated or activated in situ. Binding strength, enzyme stabilization, ease of use, protein charge 

Shape Bead, flat sheet or hollow fiber membrane, amorphous aggregate Crystal Ease of filtration, Control of diffusion path length and flow properties Simple preparation 

Surface area 50 m2/g Binding capacity, volumetric activity 

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Entrapment and Encapsulation - Biological components are physically included in a compartment. No direct chemical modifications of molecules are... 

Physical immobilization methods do not involve covalent bond formation with the enzyme, so that the native composition of the enzyme remains unaltered. Physical immobilization methods are subclassified as adsorption, entrapment, and encapsulation methods. Adsorption of proteins to the surface of a carrier is, in principle, reversible, but careful selection of the carrier material and the immobilization conditions can render desorption negligible. Entrapment of enzymes in a cross-linked polymer is accomplished by carrying out the polymerization reaction in the presence of enzyme the enzyme becomes trapped in interstitial spaces in the polymer matrix. Encapsulation of enzymes results in regions of high enzyme concentration being separated from the bulk solvent system by a semipermeable membrane, through which substrate, but not enzyme, may diffuse. Physical immobilization methods are represented in Figure 4.1 (c-e). 

The behavior of immobilized enzymes differs from that of dissolved enzymes because of the effects of the support material, or matrix, as well as conformational changes in the enzyme that result from interactions with the support and covalent modification of amino acid residues. Properties observed to change significantly upon immobilization include specific activity, pH optimum, Km, selectivity, and stability.23 Physical immobilization methods, especially entrapment and encapsulation, yield less dramatic changes in an enzyme s catalytic behavior than chemical immobilization methods or adsorption. The reason is that entrapment and encapsulation result in the enzyme remaining essentially in its native conformation, in a hydrophilic environment, with no covalent modification. 

There are commonly employed several main types of methods for immobilization of biomolecules employed in design of biosensors, including physical adsorption on the surface of transducer, covalent binding, entrapment and encapsulation, and also cross-linking. 

In general, enzyme immobilization can provide stability to bioelectrodes that ranges from minutes to hours for adsorption to months for entrapment and encapsulation. It is critical to remember that many... 

Techniques that are used to immobilize biological components for use in sensors include entrapment and encapsulation covalent binding crosslinking and absorption these various methods have been developed for proteins and there is a wide variety of chemical techniques to carry out each of these methods. 

Techniques of attachment, entrapment, and encapsulation are most widely used for cell immobilization with support materials, which are illustrated in Figure 7.1. These techniques can be applied to essentially all the viable or nonviable wholecell systems of potential interest microorganisms, plant cells, and mammalian and insect cells [2]. Although most of the principles associated with enzyme immobilization are directly applicable to cell immobilization, due to the complete difference in size and biochemical properties between enzymes the cells, the relative importance of these methods is considerably different [10]. 

One of the most important uses of calcium alginate gels is in the entrapment and encapsulation that they provide for a variety of substances such as whole cells [107], enzymes [108a], and hormones [109]. Such materials can be used in many biotechnological and biomedical applications [110]. 

Some non-silica sol-gel materials have also been developed to immobilize bioactive molecules for the construction of biosensors and to synthesize new catalysts for the functional devices. Liu et al. [33] proved that alumina sol-gel was a suitable matrix to improve the immobilization of tyrosinase for detection of trace phenols. Titania is another kind of non-silica material easily obtained from the sol-gel process [34, 35], Luckarift et al. [36] introduced a new method for enzyme immobilization in a bio-mimetic silica support. In this biosilicification process precipitation was catalyzed by the R5 peptide, the repeat unit of the silaffin, which was identified from the diatom Cylindrotheca fusiformis. During the enzyme immobilization in biosilicification the reaction mixture consisted of silicic acid (hydrolyzed tetramethyl orthosilicate) and R5 peptide and enzyme. In the process of precipitation the reaction enzyme was entrapped and nm-sized biosilica-immobilized spheres were formed. Carturan et al. [11] developed a biosil method for the encapsulation of plant and animal cells. 

The attachment and encapsulation of metals and metal complexes in the cavities of zeolites is an active area of research and provides a versatile method for the modification of these molecular sieves (39). Because of the enforced dispersion of the metal complexes in the zeolite, systems not readily observable in solution can be investigated in zeolites. For example, the mononuclear superoxo adduct of the cobalt(HI)-ammine system, [Co(NH3 )6(00-)]2+, which would be expected to dimerize in solution, could be observed entrapped in zeolite Y (40). 

One of the key factors in biosensor design is the immobilisation technique used to attach the biorecognition molecule to the transducer surface so as to render it in a stable and functional form. The challenge is to have a stable layer (or layers) of biorecognition molecules that do not desorb from the surface and that retain their activity. Entrapment or encapsulation techniques avoid the chemical changes that usually change the structure of the enzymes and modify their recognition capacity. 

In the other procedure, Rojas et al. [215] optimized the encapsulation of BLG within PLGA microparticles prepared by the multiple emulsion solvent evaporation method. The role of the pH of the external phase and the introduction of the surfactant tween 20, in the modulation of the entrapment and release of BLG from microparticles were studied. Better encapsulation of BLG was noticed on decreasing the pH of the external phase. Addition of tween 20 increased the encapsulation efficiency of BLG and considerably reduced the burst release effect. 

For many biocontrol applications, a solid matrix that could entrap or encapsulate microorganisms is advantageous. The use of a chemical reaction between sodium alginate (NaAlg) and calcium ions to make granules containing any of a wide variety of mycoherbicides was introduced in 1983 (46). 

Recent theoretical studies of Goddard et al. confirmed the encapsulation of Rose Bengal in the dendritic box [74]. A maximum number of 4-6 molecules could be entrapped and a rationale was given for the encapsulation. 

Immunization. Higher responses compared to CFA or plain antigen Method for attachment of ligands to preformed DRV s Optimization of entrapment of GO in DRV, 24% EE% obtained Preparation and performance evaluation Optimization of entrapment and freeze-drying of DRV s Explore protection by Leishmania antigen encapsulated in DRVs... 

In order to use the DRV liposomes, and/or measure the entrapment or encapsulation efficiency (or yield) they should first be separated or purified from not entrapped solute. Depending on the MW of the entrapped solute and on the final size of the DRV liposomes this can be done by centrifugation or size exclusion chromatography). 

Liposomes are characterized by a lipid bilayer structure with clearly separated hydrophilic and hydrophobic regions. Hydrophilic portions of bilayer lipids are directed towards the internal and external aqueous phases, whereas hydrophobic portions of both lipid layers are directed towards each another, forming the internal core of the membrane. A useful feature of liposomes used for drug delivery is that they allow for localization and encapsulation both water-soluble and water-insoluble substances, either together or separately. Water-soluble materials are entrapped in... 

Figure 4.1. Enzyme immobilization methods, (a) Nonpolymerizing, (b) cross-linking, (c) adsorption, (d) entrapment, and (e) encapsulation.

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