Enzymes microencapsulation

Polyamide, collodion (cellulose nitrate), ethylcellulose, cellulose acetate butyrate or silicone polymers have been used for preparation of permanent microcapsules. This method offers a double specificity due to the presence of both the enzyme and a semipermeable membrane. Moreover, it allows simultaneous immobilization of many enzymes in a single step and the surface area for contacting the substrate and the catalyst is large. The need of high protein concentration and the restriction to low molecular weight substrates are the main limitations of enzyme microencapsulation. 

FIG. 8.23 Enzyme microencapsulation. (Reproduced from Gormsen, E., Roshholm, P., and Lykke, M., in Proceedings of the 3rd World Conference on Detergents Global Perspectives, Cahn, A., Ed., AOCS Press, Champaign, IL, 1994, p. 198. With permission.)... 

Behlke M. Progress towards in vivo use of siRNAs. Mol Therapy 2006 13 645-670. Qun (Tony) Liu. Enzyme Microencapsulation and Its Application for Overcoming Multidrug Resistance in Breast Cancer Treatment, Ph.D.Thesis, University of Toronto, Toronto, Canada, 2008. 

The entrapment method for immobilization consists of the physical trapping of the active components into a film, gel, fiber, coating, or microencapsulation (see Figure 44.5). This method can be achieved by mixing an enzyme or active molecule with a polymer and then crosslinking the polymer to form a lattice structure that traps the enzyme. Microencapsulated enzymes are formed by enclosing enzymes solution within spherical semipermeable polymer membranes with controlled porosity. 

Biotechnologies Cell and enzyme microencapsulation, separation of proteins, nanofibers... 

It is sometimes possible to add properties in Hquid formulations that provide additional functions. Examples in development or in commercial use as of 1993 include microencapsulation (qv) of enzymes for protection against bleach when dispersed in a Hquid detergent addition of certain polymers to protect the enzyme after it has been added to Hquid detergents (32), or to boost activity in the final appHcation addition of surfactants or wetting agents. 

Oral pancreatic enzyme supplements are available as powders, uncoated or coated tablets, capsules, enteric-coated spheres and microspheres, or enteric-coated microtablets encased in a cellulose or gelatin capsule (Table 28-2). Microencapsulated enteric-coated products are not superior to recommended doses of conventional non-enteric-coated enzyme preparations. The quantity of active lipase delivered to the duodenum appears to be a more important determinant in pancreatic enzyme replacement therapy than the dosage form. GI side effects appear to be dose related but occur less frequently with enteric-coated products. 

Enzymes can be immobilized by matrix entrapment, by microencapsulation, by physical or ionic adsorption, by covalent binding to organic or inorganic polymer-carriers, or by whole cell immobilization (5 ). Particularly impressive is the great number of chemical reactions developed for the covalent binding of enzymes to inorganic carriers such as glass, to natural polymers such as cellulose or Sepharose, and to synthetic polymers such as nylon, polyacrylamide, and other vinyl polymers and... 

Figure 9.21. (a) Microencapsulation of enzyme molecules within a semipermeable membrane. Substrate... 

Chang TMS. Stablisation of enzymes by microencapsulation with a concentrated protein solution or by microencapsulation followed by cross-linking with glutaraldehyde. Biochem Biophys Res Commun 1971b 44 1531-1536. 

Martinek et al. (1981) also studied stabilization in systems of low water content. Several enzymes have been microencapsulated into reversed micelles formed by surfactants in apolar organic solvents (see Chapter 9). The enzymes retained their catalytic activity and substrate specificity. 

If enzymes are immobilized by copolymerization or microencapsulation, the intraparticle mass-transfer resistance can affect the rate of enzyme reaction. In order to derive an equation that shows how the mass-transfer resistance affects the effectiveness of an immobilized enzyme, let s make a series of assumptions as follows ... 

In a microencapsulation method, the encapsulate—usually an oil, flavor, enzyme, or medicinal—is emulsified in a dilute aqueous gelatin sol, a polysaccharide is added, and conditions are adjusted to favor coacervation. The encapsulate should not be truly soluble in the solvent or the cosolutes and the cosolutes should be differentially soluble in the liquid solvent. As much as 60-98% of the labile substance may be harvested by microencapsulation to yield microcapsules in the form of a free-flowing powder (Sirine, 1968). 

Perugini, R, Genta, I., Pavanetto, F., Modena, T., Maculotti, K., and Conti, B. (2002), Evaluation of enzyme stability during preparation of polylactide-co-glycolide microspheres, J. Microencapsul., 19,591-602. 

Kanawjia, S.K., Pathania, V., and Singh, S. (1992). Microencapsulation of enzymes, microorganisms and flavours and their applieations in foods. Indian Dairyman, 44(6), 280-287. 

The names of enzymes comprise two parts, the first part describes their action and the second part, -use, stands for enzyme. Alkaline protease, for instance, is an enzyme that cleaves proteins. It is present in many bacteria and fungi. Proteases are produced industrially on a large scale and are added to detergents to enhance the hydrolysis of protein-containing stains. About 80 percent of all household detergents contain proteases as microencapsulated solids (about 0.02 percent). 

In developing some of the elementary principles of the kinetics of enzyme reactions, we shall discuss an enzymatic reaction that has been suggested by Levine and LaCourse as part of a system that would reduce the size of an artificial kidney. The desired result is the production of an artificial kidney that could be worn by the patient and would incorporate a replaceable unit for the elimination of tte nitrogenous waste products such as uric acid and creatinine, In the microencapsulation scheme proposed by Levine and LaCourse, the enzyme urease would be used in tire removal of urea from ti)e bloodstream. Here, the catalytic action of urease would cause urea to decompose into ammonia and carbon dioxide. The mechanism of the reaction is believed to proceed by the following sequence of elementary reactions ... 

Bachtsi AR, Kiparissides C. An experimental investigation of enzyme release from poly(vinyl alcohol) crosslinked microspheres. ] Microencapsul 1995 12 23-35. 

Enzymes may be immobilized by encapsulation in nonpermanent (e.g., liposomes) or permanent (e.g., nylon) microcapsules. The enzyme is trapped inside by a semi-permeable membrane, where substrates and products are small enough to freely diffuse across the boundary. While nonpermanent microcapsules are useful in biochemical research, only permanent microencapsulations yield analytically useful systems, because of their mechanical stability. 

The enhanced chemiluminescense obtained with the horseradish peroxidase-H202-luminol (139) system was applied to the development of a CLD biosensor for p-iodophenol, coumaric acid (26), 2-naphthol and hydrogen peroxide. The enzyme was immobilized by microencapsulation in a sol-gel matrix. LOD for the phenolic compounds were 0.83 p,M, 15 nM and 48 nM, respectively. A remote version of the enhanced biosensor was designed by directly immobilizing the enzyme on the tip of an optical fiber. This model was used for H2O2 assay. LOD was 52.2 p,M, with RSD 4.7% (w = 4) °. A bioluminescent response was obtained for phenols with pA a > 7 in the presence of a recombinant Escherichia coli strain, DPD2540, containing a fabA luxCDABE fusion this behavior may have analytical applications. 

Supporting material for clinical analysis, cell immobilization Cell encapsulation and immobilization, immobilization of enzymes, controlled release, injectable microcapsules Microencapsulation, thermoreversible gelation Lubrication applications... 

Lee outlines three different physical methods that are commonly utilized for enzyme immobilization. Enzymes can be adsorbed physically onto a surface-active adsorbent, and adsorption is the simplest and easiest method. They can also be entrapped within a cross-linked polymer matrix. Even though the enzyme is not chemically modified during such entrapment, the enzyme can become deactivated during gel formation and enzyme leakage can be problematic. The microencapsulation technique immobilizes the enzyme within semipermeable membrane microcapsules by interfacial polymerization. All of these methods for immobilization facilitate the reuse of high-value enzymes, but they can also introduce external and internal mass-transfer resistances that must be accounted for in design and economic considerations. 

Both chemical and physical methods may be used to immobilize biocatalysts while retaining or modifying their activity, selectivity, or stability. Among the techniques used for immobilization of enzymes are physical adsorption, covalent bonding, ionic binding, chelation, cross-linking, physical entrapment, microencapsulation, and retention in permselective membrane reactors. The mode of immobilization employed for a particular application depends not only on the specific choice of enzyme and support, but also on the constraints imposed by the microenvironment associated with the application. 

For patients who are unable to swallow capsules, the contents may be emptied into applesauce, jelly, or some other nonalkatine vehicle, provided that the patient does not chew the microencapsulated beads. Side effects of pancreatic enzyme products are uncommon. Perianal irritation resembling diaper rash may occur in infants fed excess quantities of enzyme powders. Hyperuricosuria also has been reported to occur secondary to pancreatic enzyme use, apparently related to their high purine content. Proximal colonic stricture (fibrosing colonopathy) is a dose-related side effect associated with lipase doses in excess of 24,000 units/kg per day. ... 

---